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31 December 2000

    Happy festive season to you all - and good coning in 2001 :

30 December 2000

    COGNETIX Corporation: new web site:
    COGNETIX Corporation, Salt Lake City, Utah, have released their new web site. This excellent and informative resource provides details about this private company whose mission is "to discover new methods to treat human disease or injury in order to improve quality of life". The Company's most advanced product research and development efforts are based on 'conopeptides', peptides derived from the venom of Conus species of predatory sea snails (cone shells). The site provides information about the pharmacological properties of the conopeptides (summarized in a Table)and their potential disease targets (which include Epilepsy, Pain, Local Anesthesia, Neuromuscular Blockade, Neuromuscular Disorders, Demyelinating Disorders (MS, multiple sclerosis; and SCI, spinal cord injury), and Cardiovascular and Cerebrovascular Disorders (Myocardial Ischemia, Arrhythmia, and Stroke). Among the Company's products are CGX 1007 (conantokin-G) for the treatment of epilepsy (reduction of seizures, Stage I Clinical Trials began September 30, 2000); and CGX 1160 (contulakin-G), a neurotensin receptor antagonist, targeted for reduction of short-term post-operative pain. On 5 Feb, 2000 Cognetix, Inc. formed a pain management collaboration with Elan Corporation, plc. by completing a licensing and collaboration agreement to develop and commercialize Cognetix's contulakin-G (CGX-1160) for short-term management of post-operative pain using Elan's proprietary MEDIPAD Drug Delivery System. In addition, CGX-1007 (an NMDA receptor antagonist), has potential for the prevention of neuropathic pain as do a number of mu-conotoxins (sodium channel blockers) shown to be effective in pre-clinical models of pain. At the present time (Dec. 2000) Cognetix had over 20 issued or allowed patents covering the composition of conopeptide sequences from a number of Conus species and methods to treat or diagnose various central and peripheral nervous system disorders and cardiovascular conditions (see List of Conotoxin Patents).

29 December 2000

24 December 2000

22 December 2000

    Man and Mollusc Web Resource:
    Rated by Ross Mayhew as "the most complete Malacological and Conchological links assemblage on the web": - Avril's Malacological/Conchological and Internet Resources Page provides excellent links for the on-line Molluscophile community: In agreement with Ross, I heartily recommend everyone check it out and bookmark it for further use.

21 December 2000

    Xenome granted $AUD1.6 million:
    Medica Holdings Limited, Press Release, 18 December 2000 announced :Xenome granted $1.6 million to further research and development
    Extract: Medica is pleased to announce that its most recent investee company, Xenome, has been offered a $1.6 million grant by the Industry Research and Development (IR&D) Board under AusIndustry's R&D Start Program. The grant represents matching funds for a project to develop novel therapeutic compounds for pain and urological disorders. Funds are contingent on certain conditions, such as execution of an Agreement satisfactory to the IR&D Board.
    Xenome has licensed a large portfolio of novel venom-based molecules from the University of Queensland and is systematically identifying their pharmacological activity and potential as new therapeutics.
    One significant lead compound acts on a key protein in the central nervous system (CNS) which may lead to novel treatment for CNS disorders such as depression and peripheral conditions such as pain and urinary incontinence.
    Xenome intends to utilise the grant funds to optimise the pharmacological activity of the molecules and conduct preclinical testing.

13 December 2000

    Superior clinical utility suggested for AM336 relative to ziconotide:
    At the December meeting of the Australiasian Society of Clinical and Experimental Pharmacologists and Toxicologists (ASCEPT) [Abstract P 1.11, Monday 4 December, 2000], Newcastle, Australia, Dr. M.T. Smith and coworkers presented a poster entitled:
    "LACK OF CROSS-TOLERANCE BETWEEN INTRATHECAL AM336 AND INTRAVENOUS MORPHINE IN CONTRAST TO THE MARKED CROSS-TOLERANCE BETWEEN ZICONOTIDE AND MORPHINE".
    MT Smith (1), FB Ross (1), R Lewis (2), JB Kurek (3) & PJ Cabot (1), (1) School of Pharmacy, Univ of Qld, Qld, 4072, (2) Centre for Drug Design and Development, Univ of Qld, Qld 4072, (3) AMRAD Operations, Vic 3121
    Abstract: The w-conopeptides, AM336 and ziconotide are potent inhibitors of N-type calcium channels and are currently in clinical development as novel analgesic agents for the alleviation of severe pain in patients that have failed to respond adequately to strong opioids such as morphine. As many of these patients have been treated with morphine, it is important to evaluate whether cross-tolerance occurs between these conopeptides and morphine. Therefore, this study was designed to quantify the extent of cross-tolerance between intrathecal (it) AM336 and intravenous (iv) morphine relative to that for ziconotide (it) and morphine (iv) in adult male Sprague-Dawley rats with chronic inflammatory pain. Hindpaw inflammation was induced by intra-plantar injection of Freund’s Complete Adjuvant (FCA, 0.15 mL) whilst rats were under brief 3% is oflurane: 97% oxygen inhalational anaesthesia. Peptides (or vehicle) were administered via a chronically implanted polyethylene cannula inserted intrathecally between L5 and L6 and exteriorized at the base of the neck. Antinociception was quantified using the paw pressure test (PPT). On day 5 post-FCA, morphine-naďve FCA-treated rats received acute bolus doses of AM336 (0.067 nmol, it) or ziconotide (0.022, 0.059 nmol, it) in a volume of 10 mL followed by a 15 mL saline flush; antinociception was quantified for 24 h post-dosing. Rats then received a chronic infusion of morphine (10 mg/24 h) via a jugular vein cannula until rats were tolerant to its antinociceptive effects (4 days). Following a 24 h washout infusion of iv saline, rats then received a 2 nd bolus intrathecal dose of the same peptide given initially, to quantify whether the induction of morphine tolerance had altered the antinociceptive potency of subsequently administered AM336 (it) or ziconotide (it). The extent of cross-tolerance between AM336 (it) and morphine (iv) was low and not significantly different from that observed for intrathecal vehicle (150 mM lactate buffer, pH 4.5) administered before and after a chronic 7-day intrathecal infusion of vehicle. In contrast, marked cross-tolerance was found between intrathecal ziconotide and morphine. This apparently differential effect of morphine tolerance on the antinociceptive effects of subsequently administered intrathecal AM336 and ziconotide, suggests that these two conopeptides interact with different subtypes of the N-type calcium channel in the spinal cord. Furthermore, our findings suggest that AM336 may have superior clinical utility relative to ziconotide. See also Smith et al 2000 Pain. 96:119-127.

12 December 2000

10 December 2000

    3-D Structure of w-Contoxin TxVII:
    Kobayashi K, Sasaki T, Sato K, Kohno T.(2000) Three-Dimensional Solution Structure of omega-Conotoxin TxVII, an L-Type Calcium Channel Blocker. Biochemistry 39:14761-14767.
    Abstract: We determined the three-dimensional structure of omega-conotoxin TxVII, a 26-residue peptide that is an L-type calcium channel blocker, by (1)H NMR in aqueous solution. Twenty converged structures of this peptide were obtained on the basis of 411 distance constraints obtained from nuclear Overhauser effect connectivities, 20 torsion angle constraints, and 21 constraints associated with hydrogen bonds and disulfide bonds. The root-mean-square deviations about the averaged coordinates of the backbone atoms (N, C(alpha), C, and O) and all heavy atoms were 0.50 +/- 0.09 A and 0.99 +/- 0.13 A, respectively. The structure of omega-conotoxin TxVII is composed of a triple-stranded antiparallel beta-sheet and four turns. The three disulfide bonds in omega-conotoxin TxVII form the classical cystine knot motif of toxic or inhibitory polypeptides. The overall folding of omega-conotoxin TxVII is similar to those of the N-type calcium channel blockers, omega-conotoxin GVIA and MVIIA, despite the low amino acid sequence homology among them. omega-Conotoxin TxVII exposes many hydrophobic residues to a certain surface area. In contrast, omega-conotoxin GVIA and MVIIA expose basic residues in the same way as omega-conotoxin TxVII. The channel binding site of omega-conotoxin TxVII is different from those of omega-conotoxin GVIA and MVIIA, although the overall folding of these three peptides is similar. The gathered hydrophobic residues of omega-conotoxin TxVII probably interact with the hydrophobic cluster of the alpha(1) subunit of the L-type calcium channel, which consists of 13 residues located in segments 5 and 6 in domain III and in segment 6 in domain IV.

    w-Contoxins MVIIA and CVID in conscious rabbits:
    Wright, C.E., Robertson, A.D., Whorlow, S.L. and Angus, J.A. (2000) Cardiovascular and autonomic effects of omega-conotoxins MVIIA and CVID in conscious rabbits and isolated tissue assays. Br J Pharmacol. 131: 1325-1336.
    Abstract: The effects of a novel N-type voltage-operated calcium channel antagonist, omega-conotoxin CVID, were compared with omega-conotoxin MVIIA on sympathetic-evoked activation of right atria (RA), small mesenteric arteries (MA) and vasa deferentia (VD) isolated from the rat. Their effects were also compared on blood pressure and cardiovascular reflexes in conscious rabbits. The pIC(50) values for MVIIA and CVID, respectively, for inhibiting sympathetic-evoked responses were equivalent in RA (8.7 and 8.7) and VD (9.0 and 8.7); however, in MA the values were 8.4 and 7.7. The cardiac to vascular (RA/MA) potency ratios, antilog (plog RA - plog MA), for MVIIA and CVID were 2 and 10. The offset rates for CVID and MVIIA were rapid, and peptide reapplication caused rapid onset of blockade, suggesting limited desensitization. In the conscious rabbit, CVID and MVIIA (100 mug kg(-1) i.v.) caused a similar fall in blood pressure and a tachycardia that rapidly reached maximum. Both peptides decreased the vagal- and sympathetic-mediated components of the baroreflex, but had no effect on the vagal nasopharyngeal reflex. The orthostatic reflex to 90 degrees tilt was blocked by MVIIA with sustained postural hypotension for >/=90 min after administration. In contrast, CVID caused postural hypotension at 30 min which recovered rapidly. Neither CVID nor MVIIA (3 mug kg(-1) i.t.) significantly altered cardiovascular variables or autonomic reflexes. In conclusion, CVID appears to be relatively weak at inhibiting the reflex response to tilt consistent with its weaker inhibition of rat mesenteric artery constriction to perivascular nerve stimulation. This may point to subtype N-type calcium channel selectivity.

    Peptide toxins and small-cell lung carcinoma :
    Sher, E., Giovannini, F., Boot, J. and Lang, B. (2000) Peptide neurotoxins, small-cell lung carcinoma and neurological paraneoplastic syndromes. Biochimie 82: 927-936.
    Abstract: Peptide neurotoxins isolated from the venom of snakes, spiders and snails have represented invaluable tools for the identification and characterisation of membrane ion channels and receptors in vertebrate cells, including human neurons. We here report on the use of these toxins for the characterisation of membrane ion channels and receptors expressed by one of the most aggressive human cancers, small-cell lung carcinoma. This tumour shares many properties with other neuro-endocrine cell types, including the ability of firing action potentials and release hormones in a calcium-dependent manner. Toxins such as alpha-bungarotoxin and omega-conotoxins, among others, have been successfully used to characterise neuronal nicotinic receptors and voltage-dependent calcium channels, respectively, in human small-cell lung carcinoma cells. These receptors and ion channels are not only crucial for the growth of this specific tumour, but also represent autoantigens against which cancer patients build an autoimmune response. Although the aim of this autoimmune response is eventually the destruction of the cancer cells, the circulating antibodies cross-react with similar ion channels and receptors present in normal neurons or other cells, causing a number of different paraneoplastic diseases, the best characterised of which is the Lambert-Eaton myasthenic syndrome. Conotoxin-based radioimmunoassays have become an invaluable tool for the diagnosis and follow up of these paraneoplastic disorders and could represent a step forward in the early diagnosis of small-cell lung carcinoma itself.

    Ziconotide - evaluation for chronic pain:
    Jain, K.K.(2000) An evaluation of intrathecal ziconotide for the treatment of chronic pain. Expert Opin Investig Drugs 9:2403-2410.
    Abstract: Ziconotide, the synthetic form of cone snail peptide varpi-conotoxin MVIIA, is a neurone-specific N-type calcium channel blocker with an analgesic and neuroprotective effect. Intrathecal ziconotide has been recommended for approval by the FDA for the management of chronic pain. Spinally administered ziconotide produces analgesia by blocking neurotransmitter release from primary nociceptive afferents and prevents the propagation of pain signals to the brain. It has an advantage over intrathecal morphine in that there is no development of tolerance after prolonged use. Systemic toxicity is considerably reduced by administration of smaller doses intrathecally and selective delivery to the site of action in the nervous system. Nevertheless, there are neurological adverse effects due to delay in clearance of ziconotide from the neural tissues. Overall, ziconotide has a favourable risk/benefit ratio with advantages over several currently available intrathecal therapies for pain.

    See also:

    Wang, Y.X., Gao, D., Pettus, M., Phillips, C. and Bowersox, S.S. (2000) Interactions of intrathecally administered ziconotide, a selective blocker of neuronal N-type voltage-sensitive calcium channels, with morphine on nociception in rats. Pain 84:271-281.

    Atanassoff, P.G., Hartmannsgruber, M.W., Thrasher, J., Wermeling, D., Longton, W., Gaeta, R., Singh, T., Mayo, M., McGuire, D. and Luther, R.R. (2000) Ziconotide, a new N-type calcium channel blocker, administered intrathecally for acute postoperative pain.Reg Anesth Pain Med. 25:274-278

    Wang, Y.X., Pettus, M., Gao, D., Phillips, C. and Scott Bowersox S. (2000) Effects of intrathecal administration of ziconotide, a selective neuronal N-type calcium channel blocker, on mechanical allodynia and heat hyperalgesia in a rat model of postoperative pain.Pain 84: 151-158.

    Gagan Mahajan, M.D. reporting on a paper by : Fellow, UC Davis Peter G. Atanassoff, Maximillian W.B. Harmannsgruber, James Thrasher, Dan Wermeling William Longton, Raymond Gaeta, Tej Singh, Martha Nayo, Dawn McGuire, and Robert Luther, Ziconotide, A New N-Type Calcium Channel Blocker, Administered Intrathecally for Postoperative Pain . Regional Anesthesia and Pain Medicine 25 (3); 2000

    Review: N-type calcium channel blockers as analgesic agents:
    Cox, B.(2000) Calcium channel blockers and pain therapy. Curr Rev Pain 4: 488-498.

    Abstract:This review focuses on the advances in the development of N-type calcium channel blockers as analgesic agents over the last 2 years. Firstly, it highlights the clinical progress with SNX-111 (Ziconotide; Elan Pharmaceuticals, Smithfield, RI) and then secondly, it outlines the various approaches being taken by researchers to design orally active, selective, small molecule modulators without the perceived disadvantages associated with SNX-111.

1 December 2000

28 November 2000

    Venoms to Drugs 2 - Scientific Meeting
    The 2nd Venoms to Drugs meeting on Heron Island on Queensland's Great Barrier Reef will NOW be held 14th - 19th of July 2002 (see entry for March 8, 2001).

    Scientific Sessions include:

  • Ion Channel Therapeutics
  • New Pharmacologies
  • Structure//Activity Relationships
  • New discovery technologies
  • Peptides & Peptidomimetics as Drugs
  • New Targets

    Scientific Organising Committee: Professor Paul Alewood, Dr Alan Harvey, Dr Richard Lewis, Professor Andre Menez, Professor Baldomero Olivera

    Venue: Heron Island is an idyllic coral cay 60km from the Australian mainland on the Great Barrier Reef. A conference in "Heron time" offers the Great Barrier Reef at its best, sunsets on the terrace, sub-tropical ambience and casual friendliness for conference delegates.

    A web site is under development for registration for this meeting and should be available at the end of 2000. In the meantime, a downloadable pdf announcement is available and information is available from the Conference Manager, Mrs Ruth Drinkwater). To be included on the emailing list for this conference,please contact the Conference Manager, r.drinkwater@imb.uq.edu.au

26 November 2000

    Glory of India Cone:
  • The Glory of India Cone: Conus Milneedwardsi, Jousseaume, 1894 is featured among the past images on the Man and Mollusc website "This is one of the "Four Glories" of the Cone world - the other three being the famous Glory of the Seas, and the Glories of Bengal and the Atlantic... It has four forms, which are found in deep water a) from Sri Lanka to Kuwait, b) From the Red Sea to Madagascar, c) around the Mauritius and Reunion Islands, and (surprise!) d) from Ryuku Is., south of Japan. The latter three forms remain extremely rare, but trawlers are regularly dredging up the W. Indian form by the hundreds: a small specimen can be had for $10 wholesale!! Like all cones, it is a carnivore, and spears fishes for its its prey with a tiny harpoon-like projectile (a specialized form of "radula" or Gastropod-tooth) and which it uses to inject its prey with a nerve toxin. The intended victim is quickly paralyzed, after which it is swallowed whole and digested at liesure. It is not known exactly what it eats, but it probably dines on Polychaete worms, or perhaps shrimp. Cones are amoungst the fastest known hunters in the animal world, and an attack can take place in only a few milliseconds! (A milli-second is one thousanth of a second, and a cone attack can take place in 10 or less of them)".

    Dangerous Cones:
  • Collection of poisonous cones", courtesy of Paul Monfils. Featured are Conus geographus, Conus striatus, Conus textile and Conus tulipa
    Cone database:
  • An interesting biligual Japanese/English shell site featuring microshells. A model for database usage on the web. The site includes a List of common names for Cones and numerous images which when clicked on shows three views of the shell together with a length scale marker in mm. In the Shells Database enter CONIDAE. Includes among others, Conus anemone, Lamarck 1810 from Littoral in Cape York, South Australia; Conus dictator from Sri Lanka; Conus monachus, Linnaeus 1758 from Darwin, Australia Conus neptunus, Reeve 1843 from the Philippines; Conus teramachii , Kuroda 1956 off Maputo, Mozambique, and Conus vicweei, OLD 1973 from Indonesia.

25 November 2000

    Mediterranean Cone species:
    In answer to a question on CONCH-L by Bill Fenzan asking for some recent literature concerning Conus vayssierei in the Mediterranean Sea, Henk K. Mienis (E-mail: mienis@netzer.org.il & mienis@hotmail.com) replied (22 Nov 2000) that he had so far looked in vain in the well known Italian journals "La Conchiglia" and "Bollettino Malacologica", although remembers having seen some pictures in one of the journals. However, in an issue of "La Conchiglia", 23 (263): 10-15 (1992), he found an interesting article by Gabriella Raybaudi Massilia dealing with the possible presence of Conus desidiosus A. Adams, 1854 near Lampedusa Island, south of Italy. Also Geiger (2000 in Boll. Malac.) referred recently to Conus desidiosus in his treatise concerning the distribution of the living Haliotis species, because in the same area in the central Mediterranean a second Haliotis species seems to occur. Henk was forced to conclude that "the number of Conus species known to be present in the Mediterranean is still far from fixed".

    Have your say : - I invite comment on this and the related question of nomenclature of Mediterranean Conus species. To have your say, join the Cone Shells Club! on Yahoo (This club is for anyone who likes to collect cone shells), or leave a note on the Cone Shells and Conotoxins Discussion List (see link to ListBot at top of this page).

15 November 2000

8 November 2000

    Card Catalogue of World-wide Shells by S. D. Kaicher - Index of species :
    For information on the number of packs and an index to the species covered go to http://coa.acnatsci.org/conchnet/kaicher.txt. This compilation was created by Paul Callomon, Elle Scientific Publications Osaka-fu, Yao-shi, Suehiro-cho 5-2-1 581-0001 Japan, in December 1999

3 November 2000

1 November 2000

30 October 2000

    European Conus:
    The European Register of Marine Species for genus Conus, lists only two species of cone shells Conus mediteranneus and Conus vayssierei, whereas the Unitas Malacologica Check List of European Marine Mollusca (CLEMAM) lists two additional species, Conus fumigatus Hwass in Bruguičre, 1792 (R28033), and Conus papilionaceus Hwass in Bruguičre, 1792 (R27983). Why is this so ? For comments on this question see the following replies at the Cone Shells Club at Yahoo.
  • The European Register of Marine Species is a research consortium funded by the European Union under the MAST(Marine Science and Technology) programme. The area covered by the project is all the continental shelf seas of Europe, from the Canaries and Azores to Greenland and north west Russia, including the Mediterranean shelf and Baltic Seas.
    "The foundation of biodiversity research and management is correctly identifying and naming species. This Concerted Action proposal will in 24 months produce a register of marine species in Europe, linked with a bibliography of identification guides, register of taxonomic experts and locations of collections of reference specimens, and an Information Pack on European marine biodiversity (based on this project`s results). The species register is novel in its geographic extent, range of taxa covered, and that it will use the state-of-the-art World Wide Web software for publication and networking. It is anticipated that the register will become a standard reference (and technological tool) for marine biodiversity training, research and management in Europe".

  • On Thursday October 26, 2000, DRAXIS Announced that Ziconotide is to Receive Priority Review from the Canadian Drug Regulators, the Therapeutic Products Programme (TPP) of Health Canada. TPP's Priority Review Process allows for a faster review to make available promising drug products for life-threatening or severely debilitating conditions for which there are few effective therapies already on the market. Dr. Martin Barkin, President and CEO of DRAXIS Health stated, ``We are very encouraged that Ziconotide has been deemed eligible for priority review by the TPP. This process is very selectively used as evidenced by the fact that in 1999 only eight NDS submissions were selected for priority review out of a total of 84. Under the Priority Review Process the approval time for Ziconotide is expected to approximate six months, far less than the standard, which can average 18 months or more.''
20 October 2000

    Conantokins may be useful in treating Parkinson's disease.:
    Amy Adams and colleagues from the Department of Pharmacology and Toxicology, University of Utah, Salt Lake City, UT 84112-5820, USA. report that two conantokins (conantokin-G, conantokin-T(G)) potentiate L-DOPA-induced contralateral rotation in 6-hydroxydopamine-treated rats. These NMDA receptor antagonists "may be useful adjuncts in the treatment of Parkinson's disease, both in prolonging the effects of L-DOPA and reducing the side effects associated with prolonged usage and high dosing."
    Adams, A. C., Layer, R. T., Tyler McCabe,R. and Keefe, K. A. (2000) Effects of conantokins on l-3,4-dihydroxyphenylalanine-induced behavior and immediate early gene expression. European Journal of Pharmacology 404:303-313.
    Abstract: Conantokins, peptides from Conus snails, are N-methyl-d-aspartate (NMDA) receptor antagonists. NMDA receptor antagonists potentiate l-3,4-dihydroxyphenylalanine (l-DOPA)-induced rotation in 6-hydroxydopamine-treated rodents, an index of anti-Parkinsonian potential. This study examined. the effects of conantokin-G, conantokin-T(G), CGS 19755, and ifenprodil on L-DOPA-induced contralateral rotation and immediate early gene (IEG) expression in 6-hydroxydopamine-treated rats. Rats received unilateral infusions of 6-hydroxydopamine into the medial forebrain bundle. Three weeks later, rats were treated with an NMDA receptor antagonist, followed by an injection of L-DOPA. Contralateral rotations were recorded for 2 h. In addition, the expression of zif268 and c-fos were examined. Conantokin-G, conantokin-T(G), and CGS 19755 potentiated L-DOPA-induced rotation. Conantokin-G and ifenprodil had no effect on L-DOPA-induced IEG expression, whereas conantokin-T(G) and CGS 19755 attenuated expression. These data suggest that conantokins may be useful in treating Parkinson's disease. Furthermore, different NMDA receptor antagonists have distinct effects on striatal gene expression.

    Subunit interface selective toxins as probes of nicotinic acetylcholine receptor structure:
    Palmer Taylor and colleagues from the Department of Pharmacology, University of California, San Diego, La Jolla 92093, USA. report on the use of subunit-selective toxins (conotoxins, waglerins, and the alpha-neurotoxin from N. mossambica mossambica) to provide insight into the nature of the binding sites at the interface of nicotinic acetylcholine receptor subunits.
    Taylor, P., Malanz, S., Molles, B.E., Osaka, H. and Tsigelny, I. (2000) Subunit interface selective toxins as probes of nicotinic acetylcholine receptor structure. Pflugers Arch 440 (5 Suppl) R115-R117
    Abstract:The pentametric assembly of the nicotinic acetylcholine receptor with two of the five subunit interfaces serving as a ligand binding sites offers an opportunity to distinguish features on the surfaces of the subunits, and their ligand specificity characteristics. The receptor from mammalian muscle, with its circular order of homologous subunits (alphagamma alphadelta beta), assembles in a unique arrangement. The residues governing assembly can be ascertained through mutagenesis. Selectivity of certain natural toxins is sufficient to distinguish between sites at the alphagamma and alphadelta subunit interfaces. By interchanging residues on the gamma and delta subunits through mutagenesis, and ascertaining how they interact with the alpha subunit, determinants forming the binding sites can be delineated. The alpha-conotoxins show a 10,000-fold preference for the alphadelta over alphagamma subunit interface with alphaepsilon falling in between. The waglerins show a 2,000-fold preference for alphaepsilon over the alphagamma and alphadelta interfaces. Finally, the alpha-neurotoxin from N. mossambica mossambica shows a 10,000-fold preference for the alphagamma and alphadelta interfaces over alphaepsilon. Identification of interactive residues through mutagenesis, when coupled with homology modeling of domains and site-directed residue modification, has revealed important elements of receptor structure.

16 October 2000

    Conotoxin from Conus californicus targets sodium channels:
    Jon-Paul Bingham, Al Burlingame, Ed Moczydlowski and Wil Gilly from Yale, UCSF and Stanford report on a new class of sodium channel conotoxin, with new selectivity, new cys frame work, and with novel post-translational modifications. This novel conotoxin, which was isolated from the milked venom of Conus californicus, is also comparatively big.
    Bingham, J.-P., Burlingame, A., Moczydlowski, E., Gilly, W.F. (2000) A new highly selective conotoxin from Conus californicus that targets voltage-gated neuronal Na+ channels of squid. Journal of General Physiology 116:12a-13a.
    Abstract: Venoms from Conus, a carnivorous marine gastropod, are a rich source of peptide toxins that target various ion channels with highly selective subtype specificity. For example, Conus snails have provided omega-conotoxins which differentiate the various isoforms of voltage-gated calcium channels, alpha-conotoxins which differentiate between various penatameric isoforms of nicotinic acetylcholine receptor channels, and mu-conotoxins which differentiate muscle, neuronal and cardiac isoforms of voltage-gated sodium channels. In our study of the endemic Conus species, Conus californicus that is found in cooler waters of the mid coastal of region of California, we demonstrated the presence of a new highly selective sodiium channel toxin present in both the crude duct venom extract and the captive milked venom. Thus far, this novel peptide toxin was found to selectively differentiate between various species isoforms of sodium channels. It blocked the voltage-gated sodium channel in giant fiber lobe (GFL) of Pacific squid, with an absence of effect on squid calcium and potassium currents and the glutamate-activated response in GFL. The lack of an effect on sodium currents of the mollusc Aplysia californica also demonstrated its selectivity at a phylogenetic level. By analysis of the captive milked venom by reverse-phase high performance liquid chromatography (RP-HPLC), and electrospray mass spectrometry (ESMS), we observed that the active peptide was a major constituent in the hydrophobic region of the HPLC profile. However, its concentration in the crude duct venom was relatively minor. Sequential isolation and Edman characterization demonstrated a high number of post-translational modifications, a high percentage of hydrophobic amino acids, and an unusual disulfide framework distinct from other previously reported conotoxins directed against voltage-gated sodium channels (mu-, delta-, and GS-conotoxins). These features provide evidence that this toxin represents a new class of sodium channel conotoxins.

9 October 2000

26 September 2000

    Collection of beautiful, rare cone shell images: Check out this image for a beautiful collection of cone shells on the Conidae page at The Shell Store.

    Cognetix studying peptide sodium channel blockers for chronic pain:
    Source: Drug Discovery Online 9/22/2000 Cognetix Inc. (Salt Lake City, UT) has received a $98,900 Phase I Small Business Innovation Research (SBIR) grant from the National Institute of Neurological Disorders and Stroke to support the company's development of conopeptides for treating chronic pain.
    Earlier this year, Cognetix and Elan Corp. (Dublin) announced a licensing and collaborative agreement to develop and commercialize Cognetix' contulakin-G (CGX-1160) for short-term management of post-operative pain.
    Isolated from the venom of predatory marine snails, conopeptides block discrete sodium channels involved in chronic pain. Conopeptides that interact with these sodium channels should do so without inducing side effects associated with prototypical sodium channel blocking drugs.
    Chronic pain afflicts approximately 34 million people in the United States, with an annual economic impact of approximately $40 billion. The causes of chronic pain syndromes are diverse; symptoms are difficult to treat using conventional analgesics and anti-inflammatory drugs.
    In addition to pain treatments, Cognetix has research and development efforts underway in epilepsy, neuromuscular blockade, regional anesthesia, neuromuscular disorders, and cardiovascular diseases.
    For more information: Cognetix Inc., 421 Wakara Way, Suite 201, Salt Lake City, UT 84108. Tel: 801-581-0400. Fax: 801-581-9555.
    Edited by Angelo DePalma
    Managing Editor, Drug Discovery Online

17 September 2000

16 September 2000

    Tueurs sublimes dans le Pacifique [Killers of the Pacific] : This popular science article about venomous cone shells and their potential medical applications is by science writer Stephane Deligeorges and was published in the journal la Recherche (No 314, November 1998) Link here for an english translation using Babelfish (http://babelfish.altavista.com/translate.dyn).

14 September 2000

    mu-Conotoxin actions on muscle : Added the following reference on mu-contoxin action in rat muscle.
    Li, R.A., Ennis, I.L., Velez, P., Tomaselli, G.F. and Marban, E. (2000) Novel Structural Determinants of mu-Conotoxin (GIIIB) Block in Rat Skeletal Muscle mu1 Na Channels. J Biol Chem 275(36):27551-27558
    Institute of Molecular Cardiobiology, The Johns Hopkins University School of Medicine, Baltimore, Maryland 21205.
    mu-Conotoxin (mu-CTX) specifically occludes the pore of voltage-dependent Na+ channels. In the rat skeletal muscle Na+ channel (mu1), we examined the contribution of charged residues between the P loops and S6 in all four domains to mu-CTX block. Conversion of the negatively charged domain II (DII) residues Asp-762 and Glu-765 to cysteine increased the IC(50) for mu-CTX block by approximately 100-fold (wild-type = 22.3 7.0 nm; D762C = 2558 250 nm; E765C = 2020 379 nm). Restoration or reversal of charge by external modification of the cysteine-substituted channels with methanethiosulfonate reagents (methanethiosulfonate ethylsulfonate (MTSES) and methanethiosulfonate ethylammonium (MTSEA)) did not affect mu-CTX block (D762C: IC(50, MTSEA) = 2165.1 250 nm; IC(50, MTSES-) = 2753.5 456.9 nm; E765C: IC(50, MTSEA) = 2200.1 550.3 nm; IC(50, MTSES-) = 3248.1 2011.9 nm) compared with their unmodified counterparts. In contrast, the charge-conserving mutations D762E (IC(50) = 21.9 4.3 nm) and E765D (IC(50) = 22.0 7.0 nm) preserved wild-type blocking behavior, whereas the charge reversal mutants D762K (IC(50) = 4139.9 687.9 nm) and E765K (IC(50) = 4202.7 1088.0 nm) destabilized mu-CTX block even further, suggesting a prominent electrostatic component of the interactions between these DII residues and mu-CTX. Kinetic analysis of mu-CTX block reveals that the changes in toxin sensitivity are largely due to accelerated toxin dissociation (k(off)) rates with little changes in association (k(on)) rates. We conclude that the acidic residues at positions 762 and 765 are key determinants of mu-CTX block, primarily by virtue of their negative charge. The inability of the bulky MTSES or MTSEA side chain to modify mu-CTX sensitivity places steric constraints on the sites of toxin interaction. PMID: 10859326

12 September 2000

    Cone Snail Venom May Help People with Epilepsy : (Reported by Mari N. Jensen on BioMedNet)
    Investigators H. Steve White and Baldomero Olivera at the ASBMB/APET conference (June 4-8, 2000) reported that a new drug in the early stages of development for particularly intractable cases of epilepsy is the conotoxin called conantokin-G.
    Conantokin-G, a 17-amino-acid peptide isolated from the venom of Conus geographus, is a selective N-methyl-D-aspartate (NMDA) antagonist that acts on NMDA-NR2b, one of the six known NMDA receptors. NMDA receptors are glutamate receptors, excitatory receptors in the vertebrate central nervous system. Drug therapy directed at such receptors may counteract the recurrent seizures that characterize epilepsy, a disease of the central nervous system that affects more than two million Americans. One-quarter of epileptic patients worldwide are resistant to the current available therapies, which include pharmaceuticals, surgery and vagal nerve stimulation.
    The NMDA receptor is thought have an important role in acute seizures, as well as the process of epileptogenesis, the development of the disorder. So an NMDA antagonist could used treat patients at risk for developing epilepsy, in addition to those who have epilepsy, says Olivera’s colleague H. Steve White, Associate Professor of Pharmacology and Toxicology at the University of Utah.
    Other NMDA antagonists have been tried as epilepsy drugs, but the attempts were abandoned due to toxic effects. White wondered whether using a selective NMDA antagonist such as conantokin-G would control seizures without the negative effects seen in more generic NMDA antagonists.
    He’s now tested various types of rats and mice using intracerebrovascular injections of conantokin-G. Unlike other NMDA antagonists, the toxin is a broad-spectrum anticonvulsant and is effective in doses that don't produce severe side effects, he says. The companies Cognetix and Medtronics are now starting work to develop the drug, he adds.
    Michel Lazdunski, Director of the Institut de Pharmacologie Moléculaire et Cellulaire du Centre National de la Recherche Scientifique in Valbonne, France, says such a drug may cause memory problems, because glutamate is a neurotransmitter that is involved in learning and memory. Nevertheless, “I think this drug might be useful for a small subset of patients that have epileptic seizures that come all the time and who cannot accept surgery.” He adds, “I think it’s fascinating that this toxin might be used for some people.”

9 September 2000

    Tom Eichhorst's Shell Related Sites :
    Tom Eichhorst's Shell Related Internet Sites on the "Of Sea and Shore" webpage are a wonderful up-to-date resource.
    http://www.ofseaandshore.com/tomlinks.htm

    Of Sea and Shore :
    Tom Rice's Of Sea and Shore site provides information about mollusks and other marine invertebrates for shell collectors and professionals. http://www.ofseaandshore.com/

8 September 2000

    Conus nobilis victor Broderip, 1842 :
    Rich Goldberg has some very nice images on his Worldwide Conchology page of Conus nobilis victor from Indonesia.
    Conus nobilis victor Broderip, 1842 "There are many species of cone shells that exhibit considerable color and pattern variation. One of our favorites is Conus nobilis victor from eastern Flores Island, Nusa Tenggara, Indonesia. What can variability tell us about a molluscan species? A variable series of shells from one population, or a widely distributed sampling can illustrate a possible relationship between similar species, or forms. In this case, extreme forms of Conus nobilis victor from one population show how similar in appearance the subspecies comes to typical forms of Conus nobilis. Many still consider Conus victor to be a full species. Based on these preliminary observations, there is little doubt about the close relationship between Conus nobilis and Conus victor, thus the trend toward subspecific ranking for the shells from Indonesia."

    Also make sure to visit the CONIDAE page at Rich's Worldwide Conchology resource for a gallery of cone shell images.
    Worldwide Conchology - "Conchology - The Art & Science of Nature®" is a non-commercial Web resource that contains hundreds of digital images of the world's most beautiful and exotic marine, land, and freshwater shells. The digital shell image gallery has recently been expanded and improved for easier access and greater navigability. The site contains multimedia slide presentations on a number of topics, popular articles on "Shell Grading" and "Preserving Chitons", databases for searching shell names, music, and a Jigsaw PuzzleMaker© created by Jim Collins that enables you to take any shell image on the Worldwide Conchology Web site, or an image of your own, and turn it into a virtual computer jigsaw puzzle. Before you leave the site, remember to sign Richard's Guest Book and let Rich know what you think about this resource. But back to the cones ! Note that his main home page on Worldwide Conchology contains an image of Conus armiger, an uncommon deepwater Caribbean species. This particular specimen was trawled off the coast of Columbia. For clubs and societies, Rich has a professionally-produced magazine format television program entitled "Focus On Conchology" as a VHS video cassette available for rental. This program takes you around the world to meet the people, go to the places, and see the trends in Conchology. Included is an interview with the cone shell expert, Bob DaMotta and a number of beautiful Cone shells are shown during the interview. Contact Rich Goldberg for rental fee and scheduling.

2 September 2000

    Tony Swann of Wheldon & Wesley offers a summary of a talk he gave at the British Museum on Shell Books to coincide with the uploading to abebooks.com of a major collection of shell books he has just purchased. This collection can be viewed by going to the Wheldon & Wesley catalogs and clicking on FINE & RARE, or, MOLLUSCA. (included are for example Book # NTID745H24 HINTON, Alan. GUIDE TO AUSTRALIAN SHELLS. Robert Brown & Associates. Port Moresby. No Date. 77 Colour Plates Illustrating over 1,600 Individual Shells Representing 1060 Distinct Species. index; fully illustrated in col. on gloss art paper; hardcover in d/w; fine condition. Considered the most authoritative work on the subject; Book # 9036 Kohn, A.J. A CHRONOLOGICAL TAXONOMY OF CONUS, 1758-1840 Smithsonian 1992 Fine Cloth; 315p., 26 plates & color frontis Book; # 1449 Kohn, A.J. Type specimens and identity of the described species of Conus, 1758-1810 229 pp., 21 plates, 6 parts, J. Linn. Soc. Zool., 1963-81 with 4 other papers on Conus, in 1 vol., roy. 8vo, half cloth (unlettered), 1959-81 Includes the first six (of eight) parts of the series of articles on type specimens of the genus Conus published in J. Linn. Soc. Zool. In 1992 the author published a revised version of the series in book form; Book # 1582 Luther R. and Chung, N. Cowries and Cones of Hong Kong, 1975 pp. 128, sm. 8vo, wrappers, Hong Kong ; Book # 1651 Marsh, J.A. and Rippingale, O.H. Cone Shells of the World, 1964 pp. 166, with 22 coloured plates, 4to, cloth (endpapers marked), Brisbane - (also Book #5792 ; and 1st edition. Book # 000952) ; Book # 20901 Rolan Mosquera, E. Aportaciones al conoscimiento de Conus ermineus Born, 1778 (Gastropoda Conidae) estudio de los estadios juveniles. 1986 In 8 , bross., pp. 8 con 6 figs. .

29 August 2000

    Evolutionary diversity among cone shell toxin families:
    Tom Duda and Steven Palumbi from Harvard have published another interesting paper on evolutionary diversity in predatory cone snails. This time, a comparison of the conotoxin multigene families among two closely related vermivorous Conus species, Conus ebreus and Conus abbreviatus.
  • Duda, T.F. Jr, Palumbi, S.R. (2000) Evolutionary Diversification of Multigene Families: Allelic Selection of Toxins in Predatory Cone Snails. Mol. Biol. Evol. 17(9):1286–1293 Department of Organismic and Evolutionary Biology, Biological Laboratories, Harvard University.
    Abstract: In order to investigate the evolution of conotoxin multigene families among two closely related vermivorous Conus: species, we sequenced 104 four-loop conotoxin mRNAs from two individuals of Conus: ebraeus and compared these with sequences already obtained from Conus: abbreviatus. In contrast to the diversity of conotoxin sequences obtained from C. abbreviatus, only two common sequence variants were recovered from C. ebraeus. Segregation patterns of the variants in these two individuals and restriction digests of four-loop conotoxin amplification products from nine additional individuals suggest that the common variants are alleles from a single locus. These two putative alleles differ at nine positions that occur nonrandomly in the toxin-coding region of the sequences. Moreover, all substitutions are at nonsynonymous sites and are responsible for seven amino acid differences among the predicted amino acid sequences of the alleles. These results imply that conotoxin diversity is driven by strong diversifying selection and some form of frequency-dependent or overdominant selection at conotoxin loci, and they suggest that diverse conotoxin multigene families can originate from duplications at polymorphic loci. Furthermore, none of the sequences recovered from C. ebraeus appeared to be orthologs of loci from C. abbreviatus, and attempts to amplify orthologous sequences with locus-specific primers were unsuccessful among these species. These patterns suggest that venoms of closely related Conus: species may differ due to the differential expression of conotoxin loci. [PMID: 10958845]

    "In conclusion, venom diversity among Conus species is facilitated by four factors: (1) diversifying selection among loci, as revealed by the rapid evolution of conotoxin loci in C. abbreviatus (Duda and Palumbi 1999a); (2) diversifying selection within loci, as shown by the concentration of substitutions within the toxin-coding region among two alleles from C. ebraeus; (3) gene duplications, possibly of already polymorphic loci, that create a genetic architecture that can maintain a diverse set of loci; and (4) differential expression of conotoxins, as implied from the absence of identifiable expressed orthologous sequences from two closely related Conus species, C. abbreviatus and C. ebraeus."

27 August 2000

    Australian Conotoxin Research:
  • Link to current Peptide and Venom Research, specifically Studies on bioactive molecules from Australia’s venomous creatures including cone shell peptides (conotoxins), spider venoms, jumper ant venoms, and snake and scorpion venoms. This research is being conducted at the Centre for Drug Design and Development (CDDD), University of Queensland, (3-D Centre, Director Prof. Peter Andrews) under the direction of Dr. Paul Alewood, and at Xenome Ltd. by Drs. Richard Lewis and Roger Drinkwater.
    XENOME, a company within MEDICA HOLDINGS (Medica is a Pooled Development Fund specialising in investments in the biotechnology sector) is focused on the discovery of new drugs to treat disorders of the nervous system by exploiting Australia's diversity of naturally occurring venoms and toxins. Xenome, a new venture formed in January 2000 as a spin-off from the University of Queensland's Institute for Molecular Biosciences, is currently evaluating a large number of toxins and related molecules and hopes to discover novel pharmacological activities, particularly those acting on the central nervous system. It is also targeting conditions such as acute pain and stroke (See Media Press Releases for 3 July 2000 "Medica has lifted stake in Xenome to 42%" and 6 March 2000 "NEW PAIN DRUG CLINICAL TRIAL APPROVED").
    Complementary NMR studies are carried out by Prof. David Craik, CDDD.
  • Link here to the Australian research laboratories of Drs. Ray Norton, at the Biomolecular Research Institute, CSIRO, Melbourne; Prof. Jim Angus and Dr. Christine Wright, Pharmacology, Univ. Melbourne; Prof David Adams, Physiology, Univ. of Queensland; and Drs. Bruce Livett, John Down and Ken Gayler, Biochemistry and Molecular Biology, Univ. Melbourne, Australia, who are working on various aspects of conotoxin structure and function.

26 August 2000

    First account of a fatal cone shell envenomation: Alerted by 'Toto' Olivera of the book by Rumphius (1705), I went looking for it in the library and was delighted to find that THE AMBONESE CURIOSITY CABINET by GEORGIUS EVERHARDUS RUMPHIUS has recently been translated, edited, and annotated with an introduction by E.M. BEEKMAN and is published (1999) by Yale Univeristy Press. This provides the first recorded account of a fatal cone shell envenomation. The following excerpt is from Book 2, HARD SHELLFISH, pp 148-154.
      "X. Voluta pennata, these are oblong like a Roll, the head is not flat, but it protrudes like that of a Turbo, with a small red point on top, having two forms, yellow and brown; The yellow kind are called Partridges (attagenes)47 or Gold Cloth, because their entire body is painted with yellow feathers, that have black edges, almost like the feathers of that Bird: The second kind (note 48) is somewhat smaller and narrower, straight like a Roll, painted brown and white like feathers and it is called Silver Cloth; both have a narrow mouth and can stick out a little tongue, that is white, edged with red and in it is a small bone, or thorn, which will hurt you , if stung by it: The third or brown kind is different, larger and also painted brown and white, but the feathers are not arranged as orderly; they have a deathly color and are finely ribbed along the whorls: Although the Attagenata are caught and eaten daily, they are not innocent of poison, which was experienced by a slave woman on Banda, who knew that she had only held this little Whelk in her hand, which she had picked up out of the Sea, while they were pulling in a Seine net; and while she was walking to the beach, she felt a slight itching in her hand, which gradually crept up her arm and through her entire body; and so she died from it instantaneously".

23 August 2000

22 August 2000

21 August 2000

    Images of Philippine Cones:

    Novel omega conotoxins from Conus catus:

      Lewis, R.J., Nielsen, K.J., Craik, D.J., Loughnan, M.L., Adams, D.A., Sharpe, I.A., Luchian, T., Adams, D.J., Bond, T., Thomas, L., Jones, A., Matheson, J.L., Drinkwater, R., Andrews, P.R., and Alewood, P.F. (2000) Novel omega-conotoxins from Conus catus discriminate among neuronal calcium channel subtypes. J. Biol. Chem. 275: 35335-35344.
      Abstract: omega-Conotoxins selective for N-type calcium channels are useful in the management of severe pain. In an attempt to expand the therapeutic potential of this class, four new omega-conotoxins (CVIA-D) have been discovered in the venom of the piscivorous cone snail, Conus catus, using assay-guided fractionation and gene cloning. Compared to other omega-conotoxins, CVID has a novel loop 4 sequence and the highest selectivity for N-type over P/Q-type calcium channels in radioligand binding assays. CVIA-D also inhibited contractions of electrically stimulated rat vas deferens. In electrophysiological studies, CVID and MVIIA had similar potencies to inhibit current through central ({alpa}(1B-d)) and peripheral (alpha(1B-b)) splice variants of the rat N-type calcium channels when coexpressed with rat beta3 in Xenopus oocytes. However, the potency of CVID and MVIIA increased when {alpa}(1B-d) and {alpa}(1B-b) were expressed in the absence of rat b3, an effect most pronounced for CVID at {alpa}(1B-d) (540-fold), and least pronounced for MVIIA at {alpa}(1B-d) (3-fold). The novel selectivity of CVID may have therapeutic implications. (1)H NMR studies reveal that CVID possess a combination of unique structural features, including two hydrogen bonds which stabilise loop 2 and places loop 2 proximal to loop 4, creating a globular surface that is rigid and well-defined.

      Sequences and selectivity of Conotoxins CVIA-D for voltage-dependent calcium channels is shown below.

      CVIA CKSTGASCRRTSYDCCTGSCRS--GRC* N-TYPE CALCIUM CHANNEL BLOCKER
      CVIB CKGKGASCRKTMYDCCRGSCRS--GRC* N/P/Q TYPE CALCIUM CHANNELS
      CVIC CKGKGQSCSKLMYDCCTGSCSRR-GKC* N/P/Q TYPE CALCIUM CHANNELS
      CVID CKSKGAKCSKLMYDCCSGSCSGTVGRC* N-TYPE CALCIUM CHANNEL

12 August 2000

    Conformational studies on contoxins:
      Belva, H. and Lange, C. (2000) Conformational studies of omega-conotoxins using electrospray mass spectrometry. Rapid Commun Mass Spectrom 14: 1433-1439. (Spectrometrie de Masse Bio-Organique, CNRS-UMR 6014, FR11, INSERM-IFR23, UFR des sciences, Universite de Rouen, 76821 Mont Saint Aignan-Cedex, France.)
      Abstract: Omega-conotoxins MVIIA and MVIIB from Conus magus were investigated for the effect of experimental conditions on their conformations, because of the presence of three disulfide bridges in these toxins. There were no significant effects of ion-source temperature, cone voltage, pH and percentage of cosolvent. We show that charge state distributions (CSDs) observed in their electrospray mass spectra are not a true reflection of the behaviour in the bulk solution because of electrostatic effects during the ion-evaporation process in the ion source. As a result it is not possible to deduce from the observed CSDs that some basic amino acids are hidden in the core of the peptide structure. This is important in view of the complementary finding that nearly all labile hydrogens are rapidly exchanged in deuterated solvents. The mass spectrometry results can be reconciled with results of NMR experiments and molecular calculations from the literature.

    Cone snails as a source of new pharmaceuticals:

      Alan Harvey's much awaited review "Strategies for discovering drugs from previously unexplored natural products" [Research focus] has been published in Drug Discovery Today, 2000, 5: 294-300
      Here is what he has to say about Cone snails: "Another rather inaccessible source of novel chemicals is the Conus genus of marine snails. However, cone snail venoms are a particularly rich source of pharmacologically active peptides, acting on a variety of receptors and ion channels [39] . One particular conopeptide, omega-conotoxin MVIIA from Conus magus, is in Phase III clinical trials for the treatment of intractable pain; it acts as a specific blocker of N-type calcium channels in the pain fibres of the spinal cord [40] . With this successful example of a use for a conopeptide, Conus venoms can be expected to be actively searched for novel lead structures [41] . Moreover, of the 500 known species of Conus, only about a dozen have been examined in any detail, and new groups of biologically active peptide continue to be found [42]. Difficulties in collecting the snails and worries about conservation will require a molecular engineering approach to expand access to the peptide constituents of most Conus venoms. This is a challenging assignment because many of the more interesting components contain several post-translational modifications [43]".
      • 39. Olivera, B.M. et al. (1999) Speciation of cone snails and interspecific hyperdivergence of their venom peptides potential – evolutionary significance of introns. Ann. New York Acad. Sci. 870, 223–237
      • 40. Penn, R.D. and Paice, J.A. (2000) Adverse effects associated with intrathecal administration of ziconotide. Pain 85, 291–296
      • 41. Shen, G.S. et al. (2000) Conopeptides: from deadly venoms to novel therapeutics. Drug Discovery Today 5, 98–106
      • 42. Walker, C.S. et al. (1999 The T-superfamily of conotoxins. J. Biol. Chem. 274, 30664–30671
      • 43. Craig, A.G. et al. (1999) Post-translationally modified neuropeptides from Conus venoms. Eur. J. Biochem. 264, 271–275

5 August 2000

    Cone Shells in the Florida Museum:
      Search the Florida Museum Malacology Master Database for Cone Shells. When asked enter for CLASS=G, Subclass=Prosobranchia, Family=CONIDAE, Genus=Conus. To begin with, search for 10 entries in Table format. To avoid timeout, searches for larger numbers (eg. 200) are best requested as Exact Search in either Table or Report format. eg. a search for the above with Species=californicus revealed 32 entries and for Species=purpurascens, 41 entries. You can do country searches too. eg. Try entering just Genus=Conus, Country=Somalia. Compile your own list eg. Australian Cones.

4 August 2000

    References on conotoxins - added the following :
      McIntosh, J.M., Corpuz, G.O., Layer, R.T., Garrett, J.E., Wagstaff, J.D., Bulaj, G., Vyazovkina, A., Yoshikami, D., Cruz, L.J. and Olivera, B.M. (2000) Isolation and characterization of a novel Conus peptide with apparent antinociceptive activity. J Biol Chem. 275:32391-32397. [from Conus marmoreus]
    • Benie, A.J., Whitford, D., Hargittai, B., Barany, G. and Janes, R.W. (2000) Solution structure of alpha-conotoxin SI. FEBS Lett. 476(3):287-295. Link here for the solution structure, pdb=1QMW, of alpha-conotoxin S. [from Conus striatus].
    • Jacobsen, R.B., Koch, E.D., Lange-Malecki, B., Stocker, M., Verhey, J., Van Wagoner, R.M., Vyazovkina, A., Olivera, B.M. and Terlau, H. (2000) Single amino acid substitutions in {kappa}-Conotoxin PVIIA disrupt interaction with the Shaker K{super+} channel. J Biol Chem. 2000 May 18 [epub ahead of print] [from Conus purpurascens]
    • McIntosh, J.M., Gardner, S., Luo, S., Garrett, J.E. and Yoshikami, D. (2000) Conus peptides: novel probes for nicotinic acetylcholine receptor structure and function. Eur J Pharmacol. 393: 205-208.. [subpopulations of alpha3beta2beta3-containing nicotinic receptors appear to be differentially sensitive to the beta3 neuronal nicotinic receptor blocker, alpha-conotoxin MII from Conus magus]
    • Lirazan, M.B., Hooper, D., Corpuz, G.P., Ramilo, C.A., Bandyopadhyay, P., Cruz, L.J. and Olivera BM.(2000) The spasmodic peptide defines a new conotoxin superfamily. Biochemistry 39: 1583-1588. {from Conus textile. The spasmodic peptide has a novel disulfide framework and distinct signal sequence which together define a new P-superfamily of conopeptides. A cDNA encoding another member of the P-superfamily was identified from a different species, Conus gloriamaris. See entry for 12 February 2000].
    Reference on neuronal nicotinic receptor and use of alpha-conotoxins-ImI and alpha-conotoxin-MII:
    • Kaiser, S. and Wonnacott, S. (2000) alpha-Bungarotoxin-sensitive nicotinic receptors indirectly modulate [(3)H]dopamine release in rat striatal slices via glutamate release. Mol Pharmacol. 2000 Aug;58(2):312-318.

1 August 2000

31 July 2000

24 July 2000

    References on conotoxins and analogues - added the following :
    • Hill J.M., Alewood, P.F. and Craik, D.J. (2000) Conotoxin TVIIA, a novel peptide from the venom of Conus tulipa 2. Three-dimensional solution structure.Eur J Biochem.267:4649-4657.
    • Hill, J.M., Atkins, A.R., Loughnan, M.L., Jones, A., Adams, D.A., Martin, R.C., Lewis, R.J., Craik, D.J. and Alewood, P.F. (2000) Conotoxin TVIIA, a novel peptide from the venom of Conus tulipa 1. Isolation, characterization and chemical synthesis. .Eur J Biochem. 267: 4642-4648.
    • Jones, R.M. and Bulaj, G. (2000) Conotoxins - New Vistas for Peptide Therapeutics. Current Pharmaceutical Design.6: 1249-1285.REVIEW Full article available.
    • Tisa, L,S. (2000) Interaction of omega-conotoxin and the membrane calcium transport system of escherichia coli. FEMS Microbiol Lett. 188: 97-101.
    References on contryphans - added the following:
      (The contryphan family of peptides is widely distributed in venoms of the fish-hunting cone snails. Contryphan-R from Conus radiatus, contryphan-Sm from Conus stercusmuscarum and contryphan-P from Conus purpurascens venom are disulfide-constrained octapeptides containing a D-tryptophan. Contryphan-R from Conus radiatus produces the "stiff-tail" syndrome in mice).
    • Pallaghy, P.K. and Norton, R.S. (2000) The cyclic contryphan motif CPxXPXC, a robust scaffold potentially useful as an omega-conotoxin mimic. Biopolymers 54 :173-179.
    References on conantokins - added the following:
    • Williams, A.J., Dave, J.R., Phillips, J.B., Lin, Y., McCabe, R.T. and Tortella, F.C. (2000) Neuroprotective efficacy and therapeutic window of the high-affinity N-methyl-D-aspartate antagonist conantokin-G: In vitro (Primary cerebellar neurons) and In vivo (Rat model of transient focal brain ischemia) studies. J Pharmacol Exp Ther. 294: 378-386.

22 July 2000

    Cone shells on postage stamps:
    • Updated information about cone shells on postage stamps on my page Collection of Coneshell Stamps containing images of stamps in our own collection of "Cone shells from around the world" (with thanks to John Down).
    • Converted a list of Cone Shells on Stamps provided by Tom Walker for viewing on the web. For the moment I have linked images of the following cone shells to postage stamps featuring that particular species : Conus episcopatus Penhryn Cook Is $2; Conus vexillum vexillum Samoa $13; Conus textile Samoa $24; Conus textile Australia 45c; Conus litteratus Samoa $26; Conus tessulatus Samoa $50; Conus bandanus nigrescens Samoa $1; Conus gloriamaris Gilbert and Ellis Islands 10c. Tom is currently preparing a comprehensive catalogue with illustrations of every stamp. New images will be linked to this table as they become available. Tom is always happy for new collectors to contact him.

15 July 2000

8 July 2000

    Rare cones : In 1969, S. Peter Dance published his book "Rare Shells" in which he illustrated and discussed fifty shells which had an interesting and often exciting history. At one time during the past, each of those species was considered "rare" and virtually unobtainable no matter what assets that one had available. While many of the species are now common and easily obtainable for a pittance compared to their former cost, assembling a collection of all fifty, even today, poses a formidable challenge. Jacksonville Shell Club member Harry G. Lee was presented Mr. Dance's book as a gift in 1976. Now, some 22-plus years later, the Club has published his labours - 50 Rare Shells (1969) (8 of them Conus) on the web - no small task considering that even today some of the species are known to science from about a dozen specimens.
    • Included are the following 8 cones - Conus thomae Gmelin, 1791 - St. Thomas Cone; Conus cervus Lamarck, 1822 - Deer Cone; Conus crocatus Lamarck, 1810 - Saffron Cone; Conus gloriamaris Chemnitz, 1777 - Glory-of-the-seas Cone; Conus milneedwardsi Jousseaume, 1894 - Gloria-of-India Cone; Conus excelsus G. B.Sowerby III, 1908 - Illustrious Cone; Conus adamsonii Broderip, 1836 - Rhododendron Cone; and Conus dusaveli H. Adams, 1872 - Du Savel's Cone. [Mr. George Sangiouloglou (Athens, Greece) provided the images and Mr. Tom Eichhorst (Rio Rancho, New Mexico) assisted in scanning some of the species].

    Eduardo Marban and colleagues from the Institute of Molecular Cardiobiology, at Johns Hopkins University School of Medicine, Baltimore, reported (in the JBC on June 19, 2000) that negatively-charged residues in the sodium channel from rat skeletal muscle (specifically P-S6 residues D762 and E765 in domain II of the µ1-2 Na channels) are critical for high-affinity binding of µ-conotoxin GIIIB from Conus geographus. The precise localization of these residues within the pore is also crucial for optimal interactions between µ-CTX and the Na channel. "These P-S6 residues exert their effects by stabilizing the bound toxin-channel complex via electrostatic interactions with the toxin. Furthermore, the toxin may interact with the pore in certain preferred orientations, highlighting the highly asymmetrical nature of the Na+ channel pore".
    • Li, R.A., Ennis, I.L., Vélez, P., Tomaselli, G.F. and Marban, E. (2000) "Novel structural determinants of µ-conotoxin (GIIIB) block in rat skeletal muscle (µ1) Na+ channels" Journal of Biological Chemistry June 19, 2000 (M909719199)
      Abstract: mu-Conotoxin (µ-CTX) specifically occludes the pore of voltage-dependent Na+ channels. In the rat skeletal muscle Na+ channel (1), we examined the contribution of charged residues between the P loops and S6 in all four domains to µ-CTX block. Conversion of the negatively-charged domain II (DII) residues D762 and E765 to cysteine increased the IC50 for µ-CTX block by ~100-fold. Restoration or reversal of charge by external modification of the cysteine-substituted channels with methanethiosulfonate (MTS) reagents did not affect µ-CTX block relative to their unmodified counterparts. In contrast, the charge-conserving mutations D762E and E765D preserved wild-type blocking behavior while the charge-reversal mutants D762K and E765K destabilized µ-CTX block even further, suggesting a prominent electrostatic component of the interactions between these DII residues and µ-CTX. Kinetic analysis of µ-CTX block reveals that the changes in toxin sensitivity are largely due to accelerated toxin dissociation (koff) rates with little changes in association (kon) rates. We conclude that the acidic residues at positions 762 and 765 are key determinants of µ-CTX block, primarily by virtue of their negative charge. The inability of the bulky MTSES or MTSEA side chain to modify µ-CTX sensitivity places steric constraints on the site of toxin interaction.

7 July 2000

6 July 2000

    Ziconotide "approved" : [http://www.elancorp.com/news/display.asp?id=151]
    On the 28 June 2000, Elan Corporation (see also Elan Pharmaceuticals) announced receipt of an approvable letter from the U.S. Food and Drug Administration ('FDA') for Ziconotide, a novel N-type neuronal calcium channel blocker, for the treatment of severe chronic pain via the intrathecal route. This follows their submission of a New Drug Application ("NDA") 6 months ago (29 December 1999) for ziconotide as "a potential innovative new therapy for the treatment of intractable pain". For a fascinating account of the development of this novel medication see two articles in Worth On-Line magazine by Craig Canine : Pain, Profit, and Sweet Relief (Part 1) and Part 2 and 98/03-Follow-Up by Nick Pachetti entitled Neurex's Pretty Poison.

    Ziconotide, formerly known as SNX-111 was developed by Neurex Corp. in Menlo Park, California and is an N-type calcium channel blocker based on a natural component of the venom from a fish-hunting cone snail, Conus magus. The natural compound is a small peptide, an omega-conotoxin (omega-conotoxin MVIIA).

4 July 2000

    Ever since seeing the etching of a Conus marmoreus by Rembrant that was clearly sinistral (rather than the normal dextral configuration), I have been interested to know to what extent sinistral forms of Conus occur in nature. The question "does anyone know how frequent sinistral cones are in modern-day oceans?" was posed recently to the Conchologists of America List, CONCH-L by Ross Mayhew and elicited the following responses:

    From Guido T. Poppe
    My experience in Conidae C. ventricosus (mediterraneus) is rare when left handed but not impossible to obtain. We had about 6 over the latest decade. Apart from this, I've seen only two left handed Conus furvus. The sinistral Conus gloriamaris are all fakes of course.

    From Harry G. Lee
    To the references made earlier on abnormally-reversed Conus species, let me add that I have sinistral specimens of the following cones in my collection: Conus anabathrum Crosse, 1865; Conus baccatus Sowerby, 1877; Conus furvus Reeve, 1843; Conus infrenatus Reeve, 1848; Conus tinianus Hwass, 1792; and Conus ventricosus Gmelin, 1791. I hasten to add, splitting or lumping notwithstanding, Contraconus seems to be a monophyletic enterprise, and, regardless of how many trivial taxa derived, it was a lineage which prospered but lived only a few million years - a trice geologically.

2 July 2000

30 June 2000

    Mitchell Leslie, Science Writer for Stanford University Medical Center, Office of News and Public Affairs, reviewed this homepage for Science Magazine
  • SITE VISIT: When Snails Attack Science 2000 June 30; 288: 2279d (in NetWatch)
  • With news of sequencing of the human genome, added a news update page on Genetics

28 June 2000

26 June 2000

19 June 2000

Provided the following links in the Resources page:

15 June 2000

14 June 2000

    Added the following referencences on conotoxin peptide synthesis :
  • Hargittai, B. and Barany, G. (1999) Controlled syntheses of natural and disulfide-mispaired regioisomers of alpha-conotoxin SI. : J Pept Res 54: 468-479
  • Mezo, G., Drakopoulou, E., Paal, V., Rajnavolgyi, E., Vita, C. and Hudecz, F. (2000) Synthesis and immunological studies of alpha-conotoxin chimera containing an immunodominant epitope from the 268-284 region of HSV gD protein. J Pept Res 55:7-17

10 June 2000

9 June 2000

    Dr. Livett and party returned from the Field Trip at Lizard Island with a good assortment of cones (over 13 different species) and up to quota of 5 specimens in three species. Those collected in excess of GBRMA Permit quota were returned to sea to the site they were collected (ie. their own habitat). The weather was extreemly windy (min. 25 knots gusting to 60 knots)which made the small boat transfers exciting and wet. The live specimens were transported back to Melbourne in the aeroplane (in the dog and cat compartment!!) and are now living happily in our marine aquaria.

27 May 2000

    Dr. Livett is on a Field Trip at Lizard Island with 20 others until 6 June. May the cone shells be plentiful and the seas not too rough.

25 May 2000

    Latest addition to the ziconotide literature from the laboratory of George Miljanich, Elan Pharmacuticals Inc., Menlo Park, California (formerly Neurex Corp) reports on functional studies with ziconotide analogues. Ziconotide, an N-type calcium channel blocker, is the chemically synthesized form of omega conotoxin MVIIA from Conus magus.
    • Newcomb R, Abbruscato TJ, Singh T, Nadasdi L, Davis TP, Miljanich G (2000) "Bioavailability of Ziconotide in brain: influx from blood, stability, and diffusion" Peptides 21(4):491-501.
      Abstract: Ziconotide (SNX-111) is a selective peptide antagonist of the N-type calcium channel currently in clinical trials for analgesia. Ziconotide reached a maximal brain concentration of between 0.003 and 0.006% of the injected material per gram of tissue at 3-20 min after i.v. injection, and this decayed to below 0.001%/g after 2 h. The structurally distinct conopeptide SNX-185 (synthetic TVIA) was considerably more persistent in brain after i.v. administration, with 0.0035% of the injected material present at 2-4 h after i.v. injection, and 0.0015% present at 24 h. Similar results (i.e. greater persistence of SNX-185) were obtained when the peptides were perfused through in vivo dialysis probes implanted into the hippocampus. Image analysis and serial sectioning showed that diffusion of Ziconotide in the extracellular fluid around the dialysis probe was minimal, with the peptide located within 1 mm of the probe after 2 h. In vitro diffusion through cultured bovine brain microvessel endothelial cells (BBMEC) verified that a close structural analog of Ziconotide (SNX-194) passed through this blood-brain barrier (BBB) model as expected for peptides of similar physical properties (permeability coefficient of 6.5 x 10(-4) cm/g). Passage from blood to brain was also verified by in situ perfusion through the carotid artery. A statistically greater amount of radioactivity was found to cross the BBB after perfusion of radioiodinated Ziconotide compared to [(14)C]inulin. Capillary depletion experiments and HPLC analysis defined the brain location and stability. [Elan Pharmaceuticals Inc., 3760 Haven Ave. 94305, Menlo Park, CA, USA], PMID: 10822104

23 May 2000

    Updated the Resources page to include reference to:
  • Rik J Deitsch of Florida Atlantic University, who maintains the U.S.S. Conotoxin Homepage. This site provides a comprehensive list of Atlantic Cones by Dr. Edward Petuch as well as a list and directory of where certain Cone species can be found in Florida waters and the best way to collect them. In the future, a complete database of their sequences, including bioassay work with each sequence performed at Cognetix will be added.

21 May 2000

  • The Archive at the Californian Academy of Sciences covering 1994-1997, lists two articles on Cone Shells (Conus).
      Burghardt, Glenn E.(1967) "Snail with a Spear" - California Wild 20:(3) 18.
      Abstract: Description of the deadly cone Conus striatus, and how it is fed at Steinhart Aquarium.
      Nybakken, James (1974) "Carnivorous Conus" - California Wild 27: (5), 23-27.
      Abstract: Cone shells, one of the largest genera in the animal kingdom, are discussed, including their predatory behavior on other mollusks, polychaete worms, and fish, and their habitats.
    This searchable index contains entries for approximately 2000 articles from California Wild magazine (including those published under its previous title, Pacific Discovery), dating back to 1948. The full text of articles published in issues from the Summer of 1996 to the present are available online, (includes a fascinating article on tetrodotoxin) along with selected articles from 1994 and 1995. Articles prior to that date are currently available by locating back issues of the magazine at the Academy library in San Francisco or at other academic libraries. A search of this library with the keyword "Conus" yielded the following 8 entries:

    1. Crosse, Hippolyte. "Observations sur le genre Cone et description de trois especes nouvelles avec un catalogue alphabetique des Cones actuellement connus." [Paris: Bouchard-Huzard], 1858.
    2. Museum d'histoire naturelle de Geneve. "Catalogue des types et des exemplaires de Cones, figures ou decrits par Hwass, Bruguiere, Lamarck, de Lessert, Kiener et Chenu, se trouvant au Musee de Geneve." Revue suisse de zoologie; t. 54, no 5. Geneve: Impr. A. Kundig, 1947.
    3. Kohn, Alan J. "Life history and biogeography: patterns in Conus". Oxford biogeography series; 9. Oxford [England]: Clarendon Press; Oxford [England]; New York: Oxford University Press, 1994.
    4. Kohn, Alan J. A chronological taxonomy of Conus, 1758-1840. Washington: Smithsonian Institute Press, c1992.
    5. Dodge, Henry. A historical review of the mollusks of Linnaeus. New York [American Museum of Natural History] 1952-59.
    6. Marsh, J. A. Cone shells of the world. [Brisbane]: Jacaranda Press, [1964].
    7. Hanna, G Dallas, 1887-1970. West American mollusks of the genus Conus-II. Occasional papers of the California Academy of Sciences; no. 35. San Francisco, The Academy, 1963.
    8. Hanna, G Dallas, 1887-1970. West American mollusks of the genus Conus. Proceedings of the California Academy of Sciences; v. 26, no. 9. San Francisco, The Academy, 1949.

    • A brand new exhibit at the Californian Academy of Sciences, Venoms, Striking Beauties allows you to explore the world of fangs, stingers, poisonous harpoons and how they work. Lots of live animals and hands-on displays. Includes Black Widow Spider, Blue Ringed Octopus, Gila Monster, Jellyfish, Rhinocerus Viper, Death Stalker Scorpion and Velvet Ant Wasp. (Opened March 25 at the California Academy of Sciences; Golden Gate Park, San Francisco).
    • Not strictly cone shell material but nevertheless this article by Joe Slowinski will be of interest to those with an interest in venomous animals and their toxins. Striking Beauties Venomous snakes have evolved elaborate toxic cocktails to subdue and consume prey and to combat predators. It's a good idea to give them some room.

20 May 2000

14 May 2000

    The 20th Blankenese Conference on: "Animal Venoms: From Neurotoxins to Clotting Factors - Structure, Signal Transduction, Clinical Implications", was held in Hamburg-Blankenese, Germany, May 10 - 14, 2000.
    Among the invited speakers on conotoxins were 'Toto' Olivera (USA), Johan Stenflo (SWEDEN) and Mike Fainzilber (ISRAEL).

    Session 1:Conotoxin diversity (Chairperson DIETMAR RICHTER, HAMBURG) had as speakers,
    BALDOMERO OLIVERA (SALT LAKE CITY)
    Generating molecular diversity in Conus venoms
    JOHAN STENFLO (MALMÖ)
    Gamma-carboxyglutamic acid-containing conotoxins in the venom from Conus textile
    MICHAEL FAINZILBER (REHOVOT)
    Evolution and biosynthesis of conotoxins

    Short communications
    BARBARA FURIE (BOSTON)
    Vitamin K-dependent biosynthesis of gamma-carboxyglutamic acid in conotoxins from Conus textile, a venomous marine snail
    LEISA STENBERG (MALMÖ) Monoclonal antibodies specific for gamma-carboxyglutamyl (Gla) residues and their usefulness for the identification and purification of Gla-containing proteins in venom

    Posters:

  • 1. Balaji, R.A., Ohtake, A., Gopalakrishnakone, P., Sato, K., Kini, R.M., Imoto, K. & Bay, B.-H. (Singapore, Tokyo, Okazaki). Purification and partial biochemical characterization of novel vasodilatory conotoxin from Conus marmoreus venom.
  • 5. Conticello, S., Kowalsman, N. & Fainzilber, M. (Rehovot). Evolution and biosynthesis of conotoxins.
  • 6. Furie, B. & Furie, B.C. (Boston). Vitamin K-dependent biosynthesis of gamma-carboxy-glutamic acid in conotoxins from Conus textile, a venomous marine snail.
  • 19. Stenberg, L.M., Brown, M.A., Persson, U. & Stenflo, J. (Malmö). Monoclonal antibodies specific for gamma-carboxyglutamyl (Gla) residues and their usefulness for the identification and purification of Gla-containing proteins in venom.

1 May 2000

    A review by H.R. Arias from the Instituto de Matematica de Bahia Blanca, Consejo Nacional de Investigaciones Cientificas y Tecnicas (CONICET) and Universidad Nacional del Sur, Av. Alem 1253, 8000 Bahia Blanca, Argentina, addresses the question of the identification of the amino acid residues that constitute the binding sites for agonists and antagonists (such as alpha-conotoxins) on nicotinic acetylcholine receptors.

  • Arias, H.R. (2000) "Localization of agonist and competitive antagonist binding sites on nicotinic acetylcholine receptors" Neurochem. Int. 36: 595-645
    Abstract: Identification of all residues involved in the recognition and binding of cholinergic ligands (e.g. agonists, competitive antagonists, and noncompetitive agonists) is a primary objective to understand which structural components are related to the physiological function of the nicotinic acetylcholine receptor (AChR). The picture for the localization of the agonist/competitive antagonist binding sites is now clearer in the light of newer and better experimental evidence. These sites are located mainly on both alpha subunits in a pocket approximately 30-35 A above the surface membrane. Since both alpha subunits are identical, the observed high and low affinity for different ligands on the receptor is conditioned by the interaction of the alpha subunit with other non-alpha subunits. This molecular interaction takes place at the interface formed by the different subunits. For example, the high-affinity acetylcholine (ACh) binding site of the muscle-type AChR is located on the alphadelta subunit interface, whereas the low-affinity ACh binding site is located on the alphagamma subunit interface. Regarding homomeric AChRs (e.g. alpha7, alpha8, and alpha9), up to five binding sites may be located on the alphaalpha subunit interfaces. From the point of view of subunit arrangement, the gamma subunit is in between both alpha subunits and the delta subunit follows the alpha aligned in a clockwise manner from the gamma. Although some competitive antagonists such as lophotoxin and alpha-bungarotoxin bind to the same high- and low-affinity sites as ACh, other cholinergic drugs may bind with opposite specificity. For instance, the location of the high- and the low-affinity binding site for curare-related drugs as well as for agonists such as the alkaloid nicotine and the potent analgesic epibatidine (only when the AChR is in the desensitized state) is determined by the alphagamma and the alphadelta subunit interface, respectively. The case of alpha-conotoxins (alpha-CoTxs) is unique since each alpha-CoTx from different species is recognized by a specific AChR type. In addition, the specificity of alpha-CoTxs for each subunit interface is species-dependent.In general terms we may state that both alpha subunits carry the principal component for the agonist/competitive antagonist binding sites, whereas the non-alpha subunits bear the complementary component. Concerning homomeric AChRs, both the principal and the complementary component exist on the alpha subunit. The principal component on the muscle-type AChR involves three loops-forming binding domains (loops A-C). Loop A (from mouse sequence) is mainly formed by residue Y(93), loop B is molded by amino acids W(149), Y(152), and probably G(153), while loop C is shaped by residues Y(190), C(192), C(193), and Y(198). The complementary component corresponding to each non-alpha subunit probably contributes with at least four loops. More specifically, the loops at the gamma subunit are: loop D which is formed by residue K(34), loop E that is designed by W(55) and E(57), loop F which is built by a stretch of amino acids comprising L(109), S(111), C(115), I(116), and Y(117), and finally loop G that is shaped by F(172) and by the negatively-charged amino acids D(174) and E(183). The complementary component on the delta subunit, which corresponds to the high-affinity ACh binding site, is formed by homologous loops. Regarding alpha-neurotoxins, several snake and alpha-CoTxs bear specific residues that are energetically coupled with their corresponding pairs on the AChR binding site. The principal component for snake alpha-neurotoxins is located on the residue sequence alpha1W(184)-D(200), which includes loop C. In addition, amino acid sequence 55-74 from the alpha1 subunit (which includes loop E), and residues gammaL(119) (close to loop F) and gammaE(176) (close to loop G) at the low-affinity binding site, or deltaL(121) (close to the homologous region of loop G) at the high-affinity binding site, are involved in snake alpha-neurotoxin binding. The above expounded evidence indicates that each cholinergic molecule binds to specific residues which form overlapping binding sites on the AChR.Monoclonal antibodies have been of fundamental importance in the elucidation of several aspects of the biology of the AChR. Interestingly, certain antibodies partially overlap with the agonist/competitive antagonist binding sites at multiple points of contact. In this regard, a monoclonal antibody directed against the high-affinity ACh binding site (alphadelta subunit interface) induced a structural change on the AChR where the low-affinity ACh locus (alphagamma subunit interface) approached to the lipid membrane.The alpha subunits also carry the binding site for noncompetitive agonists. Noncompetitive agonists such as the acetylcholinesterase inhibitor (-)-physostigmine, the alkaloid galanthamine, and the opioid derivative codeine are molecules that weakly activate the receptor without interacting with the classical agonist binding sites. This binding site was found to be located at K(125) in an amphipathic domain of the extracellular portion of the alpha1 subunit. Interestingly, the neurotransmitter 5-hydroxytryptamine (5-HT) also binds to this site and enhances the agonist-induced ion flux activity. This suggests that 5-HT may act as an endogenous modulator (probably as co-agonist) of neuronal-type AChRs. The enhancement of the agonist-evoked currents elicited by noncompetitive agonists seems to be physiologically more important than their weak agonist properties.

    A. Menez and colleagues from the Departement d'Ingenierie et d'Etudes des Proteines, CEA, Saclay, 91191 Gif-sur-Yvette cedex, France, engineered a chimeric three-fingered curaremimetic snake toxin and demonstrated that the "third loop", normally present in the longer snake toxins is, in part, responsible for the high affinity (K(d) = 1-5 nM) of long-chain toxins for alpha7 neuronal receptors.
  • Mourier, G., Servent, D., Zinn-Justin, S. and Menez, A. (2000) "Chemical engineering of a three-fingered toxin with anti-alpha7 neuronal acetylcholine receptor activity". Protein Eng. 13 :217-225.
    Abstract: Though it possesses four disulfide bonds, the three-fingered fold is amenable to chemical synthesis using a Fmoc-based method. Thus, we synthesized a three-fingered curaremimetic toxin from snake with high yield and showed that the synthetic and native toxins have the same structural and biological properties. Both were characterized by the same 2D NMR spectra, identical high binding affinity (K(d) = 22 +/- 5 pM) for the muscular acetylcholine receptor (AChR) and identical low affinity (K(d) = 2.0 +/- 0.4 muM) for alpha7 neuronal AchR. Then, we engineered an additional loop cyclized by a fifth disulfide bond at the tip of the central finger. This loop is normally present in longer snake toxins that bind with high affinity (K(d) = 1-5 nM) to alpha7 neuronal AChR. Not only did the chimera toxin still bind with the same high affinity to the muscular AchR but also it displayed a 20-fold higher affinity (K(d) = 100 nM) for the neuronal alpha7 AchR, as compared with the parental short-chain toxin. This result demonstrates that the engineered loop contributes, at least in part, to the high affinity of long-chain toxins for alpha7 neuronal receptors. That three-fingered proteins with four or five disulfide bonds are amenable to chemical synthesis opens new perspectives for engineering new activities on this fold.

30 April 2000

Some rare and special Conus - as listed for sale by Frank Walker, Gems From The Sea, 72 Peltier Street, Marlborough, MA 01752 USA.

    Conus lucidus - Cocos Island - 54mm, F+/F++, GIANT beauty, $70.00
    Conus hunti - Barbados - 21.5, F+, from 450 feet, RARE, $250.00
    Conus sanderi - Martinique - 23mm, F+/F++, "The real deal", RARE, $350.00
    Conus kerstitchi - Mexico - 38mm, F+, another deep water rarity, $150.00
    Conus n. sp - Mexica - 35mm, F+, rare, not yet named according to my sources, $175

- and some not so rare - (from Mexico)
    Conus bartschi - 30/34mm, F+/F++, deep water dived, beauties, $25.00
    Conus bartschi - 36/39mm, F+/F++, small chips, beauties, $35.00
    Conus princeps - 50/55mm, F+/F++, dark colors and markings, beauties, $5.00
    Conus princeps - 60/68mm, F+/F++, outstanding specimens, $10.00
    Conus archon - 58/65mm, F+/F++, super patterns and colors, $12.00

27 April 2000

26 April 2000

    Lloyd Bush and Glen Hanson from Pharmacology and Toxicology University of Utah and Tyler McCabe from Cognetics, Salt Lake City, Utah, have a Short Communication showing a "Selective antagonism of nigral neuropeptide responses to methamphetamine by conantokin G, a naturally occurring conopeptide".
    Bush, L., McCabe, T. and Hanson, G.R.(2000)"Selective antagonism of nigral neuropeptide responses to methamphetamine by conantokin G, a naturally occurring conopeptide". European Journal of Pharmacology 387: 55-58
    Abstract: Some conopeptides derived from cone snails act on specific subunits of the NMDA receptor and thus, exert an influence on the dopamine system. In this study, one such conopeptide, conantokin G, was administered i.c.v. in conjunction with methamphetamine, a potent central nervous system stimulant known to cause dopamine release and changes in tissue levels of neurotensin and dynorphin A in some brain structures. Both single and multiple administrations of the conantokin G preferentially attenuated the methamphetamine-induced increases in tissue levels of these neuropeptides in the substantia nigra. Conantokin G also enhanced the behavioral effects of the methamphetamine.
    Author Keywords: Conopeptide; Conantokin G; Neurotensin; Dynorphin; Methamphetamine; NMDA receptor

25 April 2000

    Drs Yang, Hu, Li, Wu and Zhou from the National Laboratory of Biomembrane and Membrane Biotechnology, College of Life Sciences, Peking University, Beijing, People's Republic of China have studied the effects of crude venom from Conus textile on the neurophysiological responses of insect, molluscan and mammalian species. The results are interesting if unpredicable.
  • Yang D, Hu K, Li C, Wu C, Zhou P (2000) Neural electrophysiological effect of crude venom of Conus textile from the South China Sea. Toxicon 38:1607-1612.
    Abstract: In the present study, we characterized effects of the crude venom from Conus textile, a marine molluscivorous snail collected from the South China Sea, on neural electrophysiological activity in insect, molluscan and mammalian species. Our results demonstrate that the venom reversibly blocks the cholinergic synaptic transmission of cockroach Periplaneta americana central nervous system, partially blocks Na(+) currents in rat hippocampal CA(1) pyramidal neurons, and enhances the excitability and spontaneous activity of the giant neurons of garden snail Achatina fulica. PMID: 10775760

    • Dr. Richard Penn and Judith Paice from Neurosurgery at Mount Sinai School of Medicine; and the Palliative Care and Home Hospice Program, North Western Memorial Hospital, Chicago, IL, 60611, USA have logged some of the adverse effects associated with intrathecal administration of ziconotide.
      Richard D. Penn and Judith A. Paice (2000) "Adverse effects associated with the intrathecal administration of ziconotide". Pain 85, (1-2) 291-296.
      Abstract The omega-conopeptide, ziconotide, is an N-type calcium-channel blocker that has been shown to produce antinociception in animals using formalin and hot-plate tests. Initial reports of intrathecal administration of ziconotide in cancer and AIDS patients whose pain was unrelieved with opioids demonstrated analgesic efficacy. Although adverse effects were reported, these appeared to be easily managed through dose reduction or symptomatic treatment. This clinical report describes the experiences of three patients with serious adverse effects associated with intrathecal ziconotide.
      Author Keywords: Conopeptide; N-type calcium-channel blocker; Ziconotide; Neuropathic pain; Intrathecal; Side effects
    • In a related study, Drs. Ridgeway, Wallace and Gerayli from the Department of Anesthesiology, University of California, San Diego, 9500 Gilman Drive 0924, La Jolla, USA have published a paper entitled "Ziconotide for the treatment of severe spasticity after spinal cord injury" - Ridgeway, B., Wallace, M. and Gerayli, A. Pain 85: 287-289.
      Abstract: Spasticity is a major clinical manifestation of spinal cord injury and upper motor neuron syndrome.

    • see also account by Steven Vaughn who claims to be the first male in the US and Europe to have the SNX-111 (ziconotide) and experienced severe side effects to the extent that he had to be taken off the drug.
      Another account by Marti (on 12/24/99) reporting side-effects with ziconotide.
  • 24 April 2000

      Emmanuel Guillot de Suduiraut, of the Philippines has updated his Eurasia Shells website (Iconographia of the Philippine malacofauna) "to be more attractive, (seduisant), and comfortable," adding around 80 new species in the families: Buccinidae-Cancillariidae- Conidae -Costellariidae-Cypraedae-Epitoniidae-Mathildidae-Mitridae-Trividae-Trochidae-Turridae-Volutidae-Xenophoridae.
    • Among the Conidae, the following species are depicted : C.ammiralis, C.articulatus, C.aulicus aurantia, C.boeticus, C.comatosa, C.consors forma turschi, C.crocatus, C.dayriti, C.ebraeus, C.floccatus forma magdalenae, C.hopwoodi, C.kinoshitai forma tamikoae, C.legatus, C.magnificus, C.moluccensis, C.musicus (2 forms), C.mustelinus, C.neptunus forma colorvariegatus, C.omaria forma viperinus, C.otohimeae, C.pergrandis, C.proximus forma cebuensis, C.sazanka, C.spirofilis, C.suduirauti, C.tessulatus (2 forms), C.textile, C.thomae and C.vexillum. Look out for the ARCHIVE and LINKS to be added shortly.

      A collaborative study between Paul Whiteaker, Al Collins and Michael Marks at the University of Colorado, and Michael McIntosh and Siqin Luo at the University of Utah, has identified a novel nicotinic acetylcholine receptor population in mouse brain.

    • Paul Whiteaker, J. Michael McIntosh, Siqin Luo, Allan C. Collins, and Michael J. Marks (2000) 125I-alpha-Conotoxin MII Identifies a Novel Nicotinic Acetylcholine Receptor Population in Mouse Brain. Molecular Pharmacology 57 (5) 913-925.
      Abstract: alpha-Conotoxin MII (CtxMII), a peptide toxin from the venom of the predatory cone snail Conus magus, displays an unusual nicotinic pharmacology. Specific binding of a radioiodinated derivative (125I-alpha-CtxMII) was identified in brain region homogenates and tissue sections. Quantitative autoradiography indicated that 125I-alpha-CtxMII binding sites have an unique pharmacological profile and distribution in mouse brain, being largely confined to the superficial layers of the superior colliculus, nigrostriatal pathway, optic tract, olivary pretectal, and mediolateral and dorsolateral geniculate nuclei. Expression of alpha-CtxMII binding sites in the nigrostriatal pathway, combined with evidence for alpha-CtxMII-sensitivity of nicotine-induced [3H]dopamine release in rodent striatal preparations indicates that 125I-alpha-CtxMII binding nicotinic acetylcholine receptors are likely to be physiologically important. Unlabeled alpha-CtxMII potently (Kii <3 nM) competed for a subset of [3H]epibatidine binding sites in mouse brain homogenates, but weakly (IC505> 10 µM) interacted with 125I-alpha-bungarotoxin and (-)-[3H]nicotine binding sites, confirming this compound's novel nicotinic pharmacology. Quantitative autoradiography revealed that alpha-CtxMII binds with high affinity at a subset of [3H]epibatidine binding sites with relatively low cytisine affinity ("cytisine-resistant" sites), resolving [3H]epibatidine binding into three different populations, each probably corresponding to a receptor subtype. The majority population seems to correspond to that which binds nicotine and cytisine with high affinity ("cytisine-sensitive" sites). Comparison of the cytisine-resistant population's distribution with that of alpha3 subunit mRNA expression suggests that the fractions both more and less sensitive to alpha-CtxMII probably contain the alpha3 subunit, perhaps in combination with different beta subunits.

    19 April 2000

      Peter Roepstorff from Odense, Denmark together with Dario Kalume from Brazil, Johan Stenflo and Bjorn Hambe from Sweden, and Eva Czerwiec, Barbara Furie and Bruce Furie from the USA, have determined the structure of two highly modified conotoxins from Conus textile.
    • The first, conotoxin e-TxIX (1.9 kDa) is the most abundant Gla (gamma-carboxy-glutamate)-containing peptide in the venom. A venom extract (200 mg)from just 5 Conus textile snails yielded 1.1 mg of purified e-TxIX, a two loop 13 amino acid glycosylated peptide. Of the 13 amino acids, 9 were post-translationally modified (4 disulfide-bonded Cys residues, 2 Gla residues, one hydroxyproline, one bromotryptophan and a galactose/Nacetylgalactosamine glycan O-linked to the threonine residue at position 10) ! Surprisingly, the C-terminus which is amidated in most conotoxins, is present as the free acid. The three dimensional structure of e-TxIX determined by NMR was previously reported by Alan Rigby in a paper (Rigby et al 1999) describing the physiological role of this conotoxin.

    • The second, conotoxin Gla(1)-TxVI described in this paper is much larger (3.6 kDa) and provides the first description of a conotoxin containing 2 bromotryptophan residues
    • .

    • Dario E. Kalume, Johan Stenflo, Eva Czerwiec, Bjorn Hambe, Barbara C. Furie, Bruce Furie and Peter Roepstorff (2000) Structure determination of two conotoxins from Conus textile by a combination of matrix-assisted laser desorption/ionization time-of-flight and electrospray ionization mass spectrometry and biochemical methods J. Mass Spectrom. 35, 145–156.

    • Abstract: Two highly modified conotoxins from the mollusc Conus textile, e-TxIX and Gla(1)-TxVI, were characterized by matrix-assisted laser desorption / ionization and electrospray mass spectrometry and also by electrospray ionization tandem and triple mass spectrometry in combination with enzymatic cleavage and chemical modification reactions. The mass spectrometric studies allowed the confirmation of the sequence determined by Edman degradation and assignment of unidentified amino acid residues, among which bromotryptophan residues and an O-glycosylated threonine residue were observed. Methyl esterification was found necessary for the site-specific assignment of the Gla residues in the peptides. Copyright(C) 2000 John Wiley & Sons, Ltd.
      KEYWORDS conotoxin; Conus textile; bromotryptophan; matrix-assisted laser desorption/ionization mass spectrometry;electrospray ionization mass spectrometry.

    16 April 2000

      Peggy Williams from Shell Elegant, Tallevast, FL, USA, has been experimenting with a digital camera (Olympus Camedia model D340R with zoom facility) during two recent trips to Gobernadora Island, Panama, January and February, 2000. She has placed several photos of live shells (taken in an aquarium) on the web. Included are Conus archon and Conus patricius. See them and also check out dates for next year's Panama trip and other trips at www.shelltrips.com/panshells/panshells.html

    14 April 2000

      Bev Swan from Gulliver, Queensland, has kindly provided me with four scanning electron micrographs of radulae from Conus textile.

    11 April 2000

      Rob Jones* and & Grzegorz Bulaj from Cognetix Inc., Salt Lake City, UT, have just published a short review "Conus Peptides - Combinatorial Chemistry at a Cone Snail's Pace" in Current Opinion in Drug Discovery & Development, 3: (2) 141-154, March 2000.
      Abstract: Conotoxins, the disulfide rich toxin components of venomous marine cone snails, have attracted a great deal of attention in recent years because of their demonstrated pharmacological receptor subtype selectivities and distinct potential for development as new CNS therapeutics. This review highlights some of the recent advancements in this field and describes a selection of contemporary papers pertaining to regioselective strategies for the synthesis of disulfide bridges in conotoxins as well as other small constrained peptides. Primarily we have covered papers that have appeared in the scientific literature over the past 2 years. A number of comprehensive conotoxin literature reviews have appeared in recent times [1-3].
      1. Olivera, B.M., (1999) Conus venom peptides: Correlating chemistry and behavior. J. Comp. Physiol. A 185: 353-359.
      2. McIntosh, J.M., Olivera, B.M. and Cruz, L.J. (1999) Conus peptides as probes for ion channels. Methods Enzymol. 294: 605-624.
      3. Adams, D.J., Alewood, P.F., Craik, D.J., Drinkwater, R.D., Lewis, R.J. (1999) Conotoxins and their potential pharmaceutical applications. Drug Dev Res 46: 219-234.

      (*Address : Rob Jones, Ph.D, Senior Scientist, Medicinal Chemistry Unit, COGNETIX INC., 421 Wakara Way, Suite 201, Salt Lake City, UT, USA, 84108. (Email: rjones@cognetix.com))

      A large and comprehensive review entitled "Conotoxins - New Vistas in Peptide Therapeutics" will appear in Current Pharmaceutical Design later this year.

    6 April 2000

    17 March 2000

    10 March 2000
    Added a review on omega-conotoxin GVIA from Conus geographus as an analgesic and neuroprotective agent, and two recent research papers, all from Ray Norton's and Jim Angus' laboratories in Melbourne, Australia.

    • Norton, R.S., Pallaghy, P.K., Baell, J.B., Wright, C.E., Lew, M.J. and Angus, J.A. (1999) Polypeptide omega-conotoxin GVIA as a basis for new analgesic and neuroprotective agents. Drug Dev. Res. 46: 206-218.
      Abstract: The w-conotoxins are small polypeptides (of around 25 residues) cross-linked by three disulfide bonds. At least two of these, w-conotoxins GVIA and MVIIA, are potent and selective blockers of N-type voltage-gated calcium channels. Administered intravenously GVIA causes postural hypotension and blocks cardiac sympathetic and vagal reflexes, homeostatic cardiovascular effects that should normally be preserved. Administered intrathecally, MVIIA and GVIA are analgesic in acute, chronic, and neuropathic pain models, and protective following ischaemia-induced neuronal injury. We have determined the three-dimensional structure of GVIA and mapped onto that structure its calcium channel binding surface. This information is now being used in the structure-based design of truncated and stabilised peptidic analogues of GVIA and in the development of peptidomimetic analogues. This article summarizes these data and outlines the strategies being pursued in the development of low molecualr weight analogues of therapeutic applications.

    • Flinn, J.P., Pallaghy, P.K., Lew, M.J., Murphy, R., Angus, J.A. and Norton, R.S. (1999) Role of disulfide bridges in the folding, structure and biological activity of omega-conotoxin GVIA. Biochim Biophys Acta. 1434:177-90.
      Abstract: Omega-Conotoxin GVIA (GVIA), an N-type calcium channel blocker from the cone shell Conus geographus, is a 27 residue polypeptide cross-linked by three disulfide bonds. Here, we report the synthesis, structural analysis by (1)H NMR and bioassay of analogues of GVIA with disulfide bridge deletions and N- and C-terminal truncations. Two analogues that retain the crucial Lys-2 and Tyr-13 residues in loops constrained by two native disulfide bridges were synthesised using orthogonal protection of cysteine residues. In the first analogue, the Cys-15-Cys-26 disulfide bridge was deleted (by replacing the appropriate Cys residues with Ser), while in the second, this disulfide bridge and the eight C-terminal residues were deleted. No activity was detected for either analogue in a rat vas deferens assay, which measures N-type calcium channel activity in sympathetic nerve, and NMR studies showed that this was due to a gross loss of secondary and tertiary structure. Five inactive analogues that were synthesised without orthogonal protection of Cys residues as part of a previous study (Flinn et al. (1995) J. Pept. Sci. 1, 379-384) were also investigated. Three had single disulfide deletions (via Ser substitutions) and two had N- or C-terminal deletions in addition to the disulfide deletion. Peptide mapping and NMR analyses demonstrated that at least four of these analogues had non-native disulfide pairings, which presumably accounts for their lack of activity. The NMR studies also showed that all five analogues had substantially altered tertiary structures, although the backbone chemical shifts and nuclear Overhauser enhancements (NOEs) implied that native-like turn structures persisted in some of these analogues despite the non-native disulfide pairings. This work demonstrates the importance of the disulfides in omega-conotoxin folding and shows that the Cys-15-Cys-26 disulfide is essential for activity in GVIA. The NMR analyses also emphasise that backbone chemical shifts and short- and medium-range NOEs are dictated largely by local secondary structure elements and are not necessarily reliable monitors of the tertiary fold.
    • Pallaghy, P.K., Melnikova, A.P., Jimenez, E.C., Olivera, B.M. and Norton, R.S. (1999) Solution structure of contryphan-R, a naturally occurring disulfide-bridged octapeptide containing D-tryptophan: comparison with protein loops. Biochemistry 38:11553-11559.
      Abstract: Contryphan-R is a disulfide-constrained octapeptide containing a D-tryptophan that was isolated recently from venom of the cone shell Conus radiatus. The polypeptide is present in two forms in solution due to cis-trans isomerization at hydroxyproline 3. The solution structure of the major form of this unusual polypeptide, determined from NMR data, consists of a well-defined fold containing a non-hydrogen-bonded chain reversal from Gly1 to Glu5, which includes a cis-hydroxyproline and a D-Trp, and a type I beta-turn from Glu5 to Cys8. The presence of a putative salt bridge between the Glu5 carboxyl group and the N-terminal ammonium group is investigated by using various solvation models during energy minimization and is compared with the results of a pH titration. A comparison of the structure of contryphan-R with other cyclic peptide structures highlights some of the key structural determinants of these peptides and suggests that the contryphan-R fold could be exploited as a scaffold onto which unrelated protein binding surfaces could be grafted. Comparison with small disulfide-bridged loops in larger proteins shows that contryphan-R is similar to a commonly occurring loop structure found in proteins.

    8 March 2000

    • The website for "The Endemic Cones of South Africa" has been updated with photos of live animals (Includes C. pictus [all forms], C. tinianus & C. infrenatus and photos of 10 other species), visit at; 'http//pw1.netcom.com/~ejpower/sacone/conepgsaH.htm
      See also, related site on The martensi - bnerdulinus - kintoki - coelinae Group of the Genus Conus
    • Roland Anderson, Secretary,The American Malacological Society, informs me that several publications are available for sale, including bound volumes of the abstracts of recent meetings and a few copies of the venerable "How to Collect Shells" (1974), a collection of articles, 107 pp. $7.95 US, postpaid) . Details are available at the AMS website, http://erato.acnatsci.org/ams/, under "Publications".
    • Added a reference describing the purification, characterization and synthesis of an anticonvulsant peptide, conantokin R (Con-R), from the venom of the fish eating snail Conus radiatus.
        White HS, McCabe RT, Armstrong H, Donevan SD, Cruz LJ, Abogadie FC, Torres J, Rivier JE, Paarmann I, Hollmann M, Olivera BM (2000)"In vitro and in vivo characterization of conantokin-R, a selective NMDA receptor antagonist isolated from the venom of the fish-hunting snail Conus radiatus" J Pharmacol Exp Ther 292(1):425-432.
      Abstract: The purification, characterization, and synthesis of conantokin-R (Con-R), an N-methyl-D-aspartate (NMDA) receptor peptide antagonist from the venom of Conus radiatus, are described. With the use of well defined animal seizure models, Con-R was found to possess an anticonvulsant profile superior to that of ifenprodil and dizocilpine (MK-801). With voltage-clamp recording of Xenopus oocytes expressing heteromeric NMDA receptors from cloned NR1 and NR2 subunit RNAs, Con-R exhibited the following order of preference for NR2 subunits: NR2B approximately NR2A > NR2C >> NR2D. Con-R was without effect on oocytes expressing the alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA) receptor subunit GluR1 or the kainate receptor subunit GluR6. In mouse cortical neurons voltage-clamped at -60 mV, Con-R application produced a slowly developing block of inward currents evoked by 10 microM NMDA and 1 microM glycine (IC(50) = 350 nM). At 3 microM, Con-R did not affect gamma-aminobutyric acid- or kainate-evoked currents. Con-R prevented sound-induced tonic extension seizures in the Frings audiogenic seizure-susceptible mice at i.c.v. doses below toxic levels. It was also effective at nontoxic doses in CF#1 mice against tonic extension seizures induced by threshold (15 mA) and maximal (50 mA) stimulation, and it partially blocked clonic seizures induced by s.c. pentylenetetrazol. In contrast, MK-801 and ifenprodil were effective only at doses approaching (audiogenic seizures) or exceeding (electrical and pentylenetetrazol seizures) those required to produce significant behavioral impairment. These results indicate that the subtype selectivity and other properties of Con-R afford a distinct advantage over the noncompetitive NMDA antagonists MK-801 and ifenprodil. Con-R is a useful new pharmacological agent for differentiation between the anticonvulsant and toxic effects of NMDA antagonists.

    • Elan Corporation, plc and The Liposome Company Inc. have announced that they have entered into a definitive merger agreement under which Elan will acquire Liposome. "The transaction represents a strategic step for Elan into the oncology market and will be a platform from which Elan can build a presence in the oncology area. Elan is presently focused on the discovery, development and marketing of therapeutic products and services in neurology and pain management and advanced drug delivery systems....Liposome has an attractive in-market product, an experienced salesforce, an efficient manufacturing capability and a research organization that has created a range of pipeline opportunities, and it is already profitable. Liposome will complement our existing activities in oncology and oncology-related fields included in our drug delivery business and alliances, and will prospectively be of benefit to us in the marketing of ziconotide in the treatment of severe chronic cancer pain."

    4 March 2000

    • Ever wondered about the specificity of binding of the omega conotoxins MIIA (ziconotide, SNXIII) and GVIA ? Search no more - it has all been done !
      Sato, K., Raymond, C., Martin-Moutot, N., Sasaki, T., Ohtake, A., Minami, K., van Renterghem, C.V., Takahashi, M. and Seagar, M.J. (2000) Binding of six chimeric analogs of omega-conotoxin MVIIA and MVIIC to N- and P/Q-type calcium channels.Biochem. Biophys. Res. Commun. 269: 254-256.
      Abstract: Replacement of the N-terminal half of omega conotoxin MVIIC, a peptide blocker of P/Q channels, with that of omega conotoxin MVIIA significantly increased the affinity for N-type calcium channels. We examined single reverse mutations from MVIIA-type to MVIIC type in the chimeric analog named AC to identify the residues essential for subtype selectivity;. Among the four residues examined, residues in the 7th and 11th postiions were shown to be important. Among the four residues examined, residues in the 7th and 11th. The roles of these two residues were confirmed by synthesizing tow MVVIIC analogs in which Pro7 and Thr11 of MVIIC were replaced with Lys7 and leu7 of MVIIC were replaced with Lys7 and Leu11 of MVIIA, respectively.
    • The Shirshov Institute of Oceanology has one of the best discussions of Taxonomy of CONIDAE that I have seen. Ten tables of images of CONIDAE are included:
      Table 1 Plate 31
      Table 2 Plate 33
      Table 3 Plate 34
      Table 4 Plate 35
      Table 5 Plate 36
      Table 6 Plate 37
      Table 7 Plate 38
      Table 8 Plate 39
      Table 9 Plate 40
      Table 10 Plate 41
      Pay them a visit.

    3 March 2000

      A nice review article from Gregory Shen, Richard Layer and Tyler McCabe can be found in the new "Trends" journal, March 2000 issue.
    • Shen,G., Layer, R. and McCabe, T. (2000) "Conopeptides: From deadly venoms to novel therapeutics". Drug Discovery Today, 5: (3) 98-106.
      Address: Cognetix, 421 Wakara Way, Suite 201, Salt Lake City, UT 84108
      Full-text version and Adobe .pdf files available from BioMedNet after you Register (Free)

    29 February 2000

      To celebrate the launch of IMBcom and the celebration of the establishment of three new spin-off companies, Xenome Ltd, Promics Pty Ltd and Genset Pacific Pty Ltd, I have added the following information links about IMBcom and Xenome to the Resources Page. Xenome Ltd. has been set up to discover and develop new drugs and drug leads from venoms and toxins of Australian aquatic and land creatures, particularly marine cone shell species. [Professor John Hay, Vice-Chancellor, The University of Queensland, will launch "IMBcom" the commercialisation arm for the The Institute for Molecular Bioscience on Thursday 9 March, 2000, 10.00 am - 11.30 am at the Art Gallery Annexe, Brisbane Customs House]

    • University of Queensland - News Releases on Xenome
    • Xenome: Snail's poison provides next-generation painkilling drugs (Dr. Richard Lewis with cone shells)
    • University research centre forms spinoff drug commercialisation companies: Xenome, Promics, Genset
    • Cone shell venom relieves pain - extract from Australian Pharmacist 19: (1), Jan/Feb 2000, p 5.
      • "[CLINICAL trials with cone shell venom at the University of Queensland may lead to a new treatment for chronic, intractable pain.
      • Researches from the University of Queensland's Centre for Drug Design and Development in collaboration with Australian pharmaceutical company, AMRAD Operations, have isolated tiny proteins or peptides from the venom of 50 species of Great Barrier Reef cone shell snails.
      • The venom has been purified, sequenced and synthesized with the extracted peptides shown to have great potential as highly selective pain-killing drugs.
      • The General Manager of Xenome Ltd, a University company established in 1998 to develop the centre's research, Roger Drinkwater says the peptide is one of the first cone peptides to be developed to this stage.
      • The team hopes the trials may provide an alternative for cancer patients for whom morphine is ineffective.
      • Work on conotoxins as a source for drugs was started in Australia 20 years ago by the late Professor Bob Endean of the university's Zoology Department who identified the potential of cone shell snail venom for pharmaceuticals.
      • Since then, the 3-D Centre's Venom Research Group led by Professor Peter Andrews, Associate Professor Paul Alewood and Dr. Richard Lewis have isolated potential peptides for new drugs.
      • 'Some of the most exciting conotoxin work by blocking specific calcium channels within nerve cells', Dr. Lewis said.
      • 'These channels are missed by current pain-relieving drugs with the effect that relief wears off quickly.'
      • The research forms part of the University's new Insititute for Molecular Bioscience, the centrepiece of a $100 million research complex being developed by the university in conjunction with CSIRO and other agencies.
      Cone shell snails range in length from 15 cm and produce venom in a duct that delivers the toxins through a proboscis in their mouths. Prey such as fish, worms or molluscs are harpooned and injected with the lethal cocktail during nightly hunting missions.
      Research staff "milked" between 50 and 100 cone shell snails to get enough venom for their initial experiments. Medica Holdings has bought 31% of Xenome Ltd.
      Photos: (1) Milking a cone shell of venom. (2) Researcher Dr. Richard Lewis with a collection of cone shells]"
    18 February 2000
      Added reference to a review (in Russian) by Drs. Utkin and colleagues (utkin@ibch.siobc.ras.ru) on contoxins from three researchers at the Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Acadamy of Sciences, Moscow.
    • Utkin, Iu.N., Kasheverov I,E. and Tsetlin, V.I. (1999) [alpha-Neurotoxins and alpha-conotoxins--nicotinic cholinoreceptor blockers]. Bioorg Khim 25: 805-810.
      Abstract: The review is devoted to the competitive blockers of different nicotinic acetylcholine receptors, alpha-neurotoxins from snake venoms, and alpha-conotoxins from marine snails of the Conidae family. The relationship between the structure and function of these toxins is discussed. Recent data on the mechanism of alpha-neurotoxin and alpha-conotoxin interaction with the nicotinic acetylcholine receptor are presented. [PMID: 10645484, UI: 20110079]
      CLUB CONCHYLIA e.V. is a club that unites collectors and scientifically minded people in the area of malacozoology (science of mollusks), and contains Links to other mollusc sites .
    • The club was founded in 1969 and currently has 380 members (from Germany, Austria, Switzerland, and other countries). The interests of the members are varied. Most collect shells of marine mollusks, some are interested in terrestrial and/or freshwater mollusks, others are involved with fossils, others specialize in stamps featuring mollusks. In addition to scientific interests, many collectors are concerned with aesthetics.
    • The diversity in interests is reflected in the Club's journals Club Conchylia Informationen and Acta Conchyliorum. In addition to scientific contributions in the areas of taxonomy, evolution, biology, and zoogeography, the journals also contain travel accounts, ethnological essays, and philatelistic contributions, as well as reports about exhibitions, fairs and meetings.
    • Each autumn, a annual convention (Jahreshauptversammlung, JHV) takes place, which contains opportunities for exchange and purchase of shells. The annual convention is organized by members and is located every year at a different place. Additionally, regional meetings are held in various areas. The club does not engage in any kind of commercial activity. The entire work of the club including the editorial duties are carried out in an honourary capacity.
    13 February 2000
      Added reference to a paper on the competition of alpha-conotoxin PnIB (from Conus pennaceus) with 125I-alpha-bungarotoxin for binding to a high-affinity construct of the neuronal alpha7 acetylcholine receptor expressed in 293 HEK cells:
    • Quiram, P.A., McIntosh, J.M. and Sine, S.M. (2000) Pairwise interactions between neuronal alpha7 acetylcholine receptors and alpha-conotoxin PnIB
      J. Biol. Chem. 275: 4889-4896
      .
      Abstract: This work uses alpha-conotoxin PnIB to probe the agonist binding site of neuronal alpha7 acetylcholine receptors. We mutated the 13 non-cysteines in CTx PnIB, expressed alpha7/ 5-hydroxytryptamine-3 homomeric receptors in 293 HEK cells, and measured binding of each mutant toxin to the expressed receptors by competition against the initial rate of 125I--bungarotoxin binding. The results reveal that residues Ser-4, Leu-5, Pro-6, Pro-7, Ala-9, and Leu-10 endow CTx PnIB with affinity for alpha7/5-hydroxytryptamine-3 receptors; side chains of these residues cluster in a localized region within the three-dimensional structure of CTx PnIB. We next mutated key residues in the seven loops of alpha77 that converge at subunit interfaces to form the agonist binding site. The results reveal predominant contributions by residues Trp-149 and Tyr-93 in alpha7 and smaller contributions by Ser-34, Arg-186, Tyr-188, and Tyr-195. To identify pairwise interactions that stabilize the receptor-conotoxin complex, we measured binding of receptor and toxin mutations and analyzed the results by double mutant cycles. The results reveal a single dominant interaction between Leu-10 of CTx PnIB and Trp-149 of alpha7 that anchors the toxin to the binding site. We also find weaker interactions between Pro-6 of CTx PnIB and Trp-149 and between both Pro-6 and Pro-7 and Tyr-93 of alpha7. The overall results demonstrate that a localized hydrophobic region in CTx PnIB interacts with conserved aromatic residues on one of the two faces of the alpha7 binding site.
    12 February 2000
      Machiko Yamada, an amateur collector from Miyagi, Japan, has an attractive and informative bilingual Japanese/English database with images and information about the following 58 members of CONIDAE. You can search on Shape, Species Name or Japanese Name. When you access the page, clicking on the small cone shell image will give you a larger image. Well worth a visit.
    • Conus caracteristicus Characteristic Cone , Conus tessulatus Tessellate Cone, Conus pulcher stamensis , Conus imperialis Imperial Cone, Conus ebreus Hebrew Cone, Conus taeniatus Ringed Cone, Conus abbreviatus Abbreviated Cone, Conus musicus Music Cone, Conus ventricosus Mediterrananean Cone, Conus arenatus Sand-dusted Cone, Conus cocceus Cocceus Cone, Conus anemone Anemone Cone, Conus ammiralis Admiral Cone, Conus nobilus Nobel Cone, Conus spurius lorenzianis Lorenz's Cone, Conus amadus Amadis Cone, Conus planorbis Planorbis Cone, Conus striatellus Striatellus Cone, Conus magus Magus Cone, Conus suturatus Violet-base Cone, Conus lemniscatus Ribbon Cone, Conus meracator Trader Cone, Conus lynceus Lynceus Cone, Conus capitaneus Captain Cone, Conus capitanaleus Little Captain Cone, Conus rattus Rat Cone, Conus pertusus Pertusus Cone, Conus flavidus Yellow Pacific Cone, Conus frigidus Frigid Cone, Conus emaciatus False Virgin Cone, Conus lividus Livid Cone, Conus floridulus , Conus pennaceus Feathered Cone, Conus praelatus , Conus legatus , Ambassador Cone, Conus gloriamaris Glory of the Sea Cone, Conus victoriae Queen Victoria Cone, Conus alulicus Princely Cone, Conus betulinus Beech Cone, Conus genuanus Garter Cone, Conus patricius Pear-shaped Cone, Conus striatus Striate Cone, Conus gubernator Governor Cone, Conus bullatus Bubble Cone, Conus coccineus Scarlet Cone, Conus glans Acorn Cone, Conus australis Austral Cone, Conus comatosa Comatose Cone, Conus cuvieri Cuvier's Cone, Conus sieboldii Siebold's Cone, Conus ichinoseana Ichinose Cone, Conus stimpsoni Stimpson's Cone, Conus kimioi Kimio's Cone, Conus eugrammatus Eugrammatus Cone, Conus recurvus Recurved Cone, Conus urashimanus Urashima Cone, Conus nux , Conus doreesis Pontifical Cone

    • Added reference to a paper from 'Toto' Olivera's laboratory describing the biological activity and molecular structure of an unusual peptide from Conus textile venom that elicits distinct behavioral symptoms in mice. Upon injection of this so-called "spasmodic peptide", normal mice assume the phenotype of a well-known mutant, the spasmodic mouse. This peptide, and a related peptide from another molluscivorous species, Conus gloriamaris, share a distinctive Cys pattern, one not previously seen in any conotoxin family (in the Conus gloriamaris peptide the two gamma-carboxyglutamates of the spasmodic peptide of Conus textile are mutated to serine and alanine (see below). Thus, the spasmodic peptide is the first member of a novel superfamily of conotoxins, which have been termed the P-superfamily. As is true of other conotoxin superfamilies, the P-superfamily can be recognized by both a unique signal sequence and distinct arrangement of Cys residues.
      • Lirazan, M.B., Hooper, D., Corpuz, G.P., Ramilo, C.A., Bandyopadhyay, P., Cruz, L.F. and Olivera, B.M. (2000) The spasmodic peptide defines a new conotoxin superfamily.
        Biochemistry . Biochemistry 39: 1583-1588.
        Abstract: We purified and characterized a peptide from the venom of Conus textile that makes normal mice assume the phenotype of a well-known mutant, the spasmodic mouse. This "spasmodic" peptide has 27 amino acids, including two gamma-carboyglutamate (Gla) residues. A cDNA clone encoding the precursor for the peptide was identified: a gamma-carboxylation recognition signal sequence (gamma-CRS) is present in the -> -1 -> -20 region of the peptide precursor. Both the gamma-CRS and the postiion of the Gla residues in the mature toxin are notably different from other Gla-containing conopeptides. The spasmodic peptide has a novel disulfide framework and distinct signal sequence which together define a new P-superfamily of conopeptides. A cDNA encoding another member of the P-superfamily was identified from a different species, Conus gloriamaris.

      The sequence of the spasmodic peptide is given as :

           5    10   15   20   25            

      GCNNSCQgHSDCgSHCICTFRGCGAVN-NH2       (tx9a)Conus textile

      SCNNSCQSHSDCASHCICTFRGCGAVN-NH2       Conus gloriamaris

      (where g is gamma-carboxyglutamate)

      A note on nomenclature: In accordance with the nomenclature of McIntosh et al 1999 (Ann.Rev. Biochem.68: 59-88 1999), all peptides with six non-adjacent Cys residues (C--C--C--C--C--C) have been designated as Cys framework 9, all peptides with this framework have '9' or 'IX' (the latter when the mechanism has been determined). Thus, the spasmodic peptide (from C.textile venom) is technically referred to as peptide tx9a, encoded by Tx9.1. When the mechanism is known, it might be renamed by a designation such as "pi-conotoxin IXA". The cDNA clone from C. gloriamaris is Gm9.1 (see Footnote to Table 1, Lirazan et al 2000).

      Summary of Cys frameworks

      P-Superfamily C--C--C---C--C--C

      O-Superfamily C--C--CC--C--C

      T-Superfamily CC---CC

      See related papers:
      1 : Craig, A.G., Speir, J.P., Rosamilia, S., Fursey, V., Lirazan, M.B. (2000) [M + Fe - 5H]2- peptide ion composition verified by Fourier transform mass spectrometry accurate mass and tandem mass spectrometry analyses. J Am Soc Mass Spectrom. 11: 83-87.

      2 : Walker, C.S., Steel, D., Jacobsen, R.B., Lirazan, M.B., Cruz, L.J., Hooper, D., Shetty, R., DelaCruz, R.C., Nielsen, J.S., Zhou, L.M., Bandyopadhyay, P., Craig, A.G., Olivera, B.M. (1999) The T-superfamily of conotoxins.J Biol Chem. 274: 30664-30671.

    31 January 2000

      Added reference to a paper from Michael McIntosh's laboratory on the effects of alpha-conotoxins MII (from Conus magus) and AuIB (from Conus aulicus) on nicotine-evoked noradrenaline release from the hippocampus of the rat:

    • Fu, Y., Matta, S.G., McIntosh, J.M. and Sharp, B.M. (1999) Inhibition of nicotine-induced hippocampal norepinephrine release in rats by alpha-conotoxins MII and AuIB microinjected into the locus coeruleus
      Neuroscience Letters 266: 113-116
      .
      Abstract: Hippocampal norepinephrine (NE) is secreted by neurons projecting from the locus coeruleus (LC) to the hippocampus; LC nicotinic receptors (NAchRs) are involved in the effects of systemic nicotine on this pathway. To clarify the NAchR subtypes, NAchR antagonists, termed alpha-conotoxins, were microinjected into the LC before nicotine; MII and AulB were used to assess the potential involvement of alpha3beta2 and alpha3beta4 subunit-containing NAchRs, respectively. dose-dependently stimulated hippocampal NE release (P<0.01). MII (>0.25 pmol) reduced the NE response to nicotine (67% decrease; P<0.05), as did AuIB (44% reduction by 25 pmol; P<0.05). Administered together, however, MII and AuIB were no more effective than MII. Thus, MII and AuIB are capable of interacting with NAchR subtypes other than those previously defined as alpha3beta2 and alpha3beta4, respectively. nAChRs containing both beta2 and beta4subunits may be involved.
      Key words: alpha-conotoxin MII; alpha-conotoxin AuIB, norepinephrine, hippocampus, locus coeruleus, nicotinic receptors, In vivo microdialysis
    • Haghighi, A.P. and Cooper, E. (2000) A molecular link between inward rectification and calcium permeability of neuronal nicotinic acetylcholine alpha3beta4 and alpha4beta2 receptors.
      J.Neurosci. 20: 529-541.
      .
      Abstract: Many nicotinic acetylcholine receptors (nAChRs) expressed by central neurons are located at presynaptic nerve terminals. These receptors have high calcium permeability and exhibit strong inward rectification, two important physiological features that enable them to facilitate transmitter release. Previously, we showed that intracellular polyamines act as gating molecules to block neuronal nAChRs in a voltage-dependent manner, leading to inward rectification. Our goal is to identify the structural determinants that underlie the block by intracellular polyamines and govern calcium permeability of neuronal nAChRs. We hypothesize that two ring-like collections of negatively charged amino acids (cytoplasmic and intermediate rings) near the intracellular mouth of the pore mediate the interaction with intracellular polyamines and also influence calcium permeability. Using site-directed mutagenesis and electrophysiology on alpha4beta2 and alpha3beta4 receptors expressed in Xenopus oocytes, we observed that removing the five negative charges of the cytoplasmic ring had little effect on either inward rectification or calcium permeability. However, partial removal of negative charges of the intermediate ring diminished the high-affinity, voltage-dependent interaction between intracellular polyamines and the receptor, abolishing inward rectification. In addition, these nonrectifying mutant receptors showed a drastic reduction in calcium permeability. Our results indicate that the negatively charged glutamic acid residues at the intermediate ring form both a high-affinity binding site for intracellular polyamines and a selectivity filter for inflowing calcium ions; that is, a common site links inward rectification and calcium permeability of neuronal nAChRs. Physiologically, this molecular mechanism provides insight into how presynaptic nAChRs act to influence transmitter release.
      Key words: nicotinic acetylcholine receptor; presynaptic receptors; transmitter release; ion permeation; gating particles
    26 January 2000 (Australia Day !)


      Emmanuel Guillot de Suduiraut (P.O. 184 Lapu-Lapu City, 6015 CEBU Philippines) has a very nice site titled Eurasia Shells at http://www.eurasiashells.freeservers.com/main.html where more than 220 species are illustrated, including several Holotypes and Paratypes. You can find unascribed specimens and others rarely represented in the literature.

      All gastropods illustrated come exclusively from Philippines and in particularly from five geographic areas well delimited. Some 38 species of Conidae are illustrated.

    25 January 2000

      MEETING ANNOUNCEMENT: 3rd International Workshop in Malacology -
      Menfi (Sicily, Italy) June 15-19, 2000.
      "Systematics, Phylogeny and Biology of Neogastropoda"
      go to its web-announcement at http://www.futuralink.it/vannarotolo/istituzione/index.htm
      1st Announcement and Registration Form

      Preliminary list of contributions on Conus: (other contributions not listed here)
      .

    • THE EVOLUTION OF THE RADULA OF CONIDAE AND TURRIDAE. E. Rolan
    • THE VENOMOUS CONE SNAILS Biology, Toxinology and Phylogeny Baldomero Olivera
    • A TALE OF REDUCTION AND LOSS FOREGUT ANATOMY OF RAPHITOMINE GASTROPODS (CONOIDEA RAPHITOMINAE). Y.I. Kantor & J.D. Taylor
    • MORPHOLOGICAL PREREQUISITES FOR UNDERSTANDING NEOGASTROPOD PHYLOGENY. Y.I. Kantor
    • FOREGUT ONTOGENY OF THE NEOGASTROPODA COMPARISON OF DEVELOPMENT IN NUCELLA AND CONUS. A.D. Ball
    • Facilities for students available !
      Enquire from:
      Marco Oliverio - Evolutionary Biology PhD
      Research Scientist
      Dipartimento di Biologia Animale e dell'Uomo
      Viale dell'Universita' 32
      I-00185 Roma ITALY
      phone +39.6.49914750
      +39.6.55176356 (Molecular Systematics lab.)
      FAX +39.6.4958259
      -----------------------------------------------------------
      Visit Italian Malacological Society Web Site at http://aicon.com/sim
      -----------------------------------------------------------


    24 January 2000

    21 January 2000

    • New publication from Mike Fainzilber's group raises interesting questions about the significance of codon conservation for cysteine in crucial residues in hypervariable gene families such as the conotoxin superfamily in invertebrates and the olfactory receptor superfamily in mammals. To estimate the possible range of variability of conopeptides, Conticello et al examined the conopeptide precursor cDNAs currently available in GenBank. The largest conopeptide family (the so-called scaffold VI/VII grouping) was chosen for analysis. The resulting multiple sequence alignment consisted of 53 conopeptide precursors from 9 different species. Examination of the corresponding nucleotide alignments showed that 5 out of 6 cysteines in these peptides exhibit a pronounced position-specific codon conservation, notably in the most hypervariable region of the sequence.
      Conticello, S. G., Pilpel, Y., Glusman, G. and Fainzilber, M. (2000) "Position-specific codon conservation in hypervariable gene families." Trends in Genetics 16: 57-59.
      Abstract:This paper addresses the question concerning conservation of the third nucleotide (T or C) in two codons (TGT or TGC) that code for cysteine. Analysis of cysteine codon usage in a number of hypervariable gene families revealed strict codon conservation in specific positions adjacent to or within the hypervariable regions of these genes. This phenomenon suggests the possible existence of specific positional codon-conservation mechanisms in certain genes. The phenomenon of position-specific codon conservation may provide a functional-genomics aproach to identify important residues for further structural and functional study. Moreover, this observation suggests that novel mechanisms could exist to conserve crucial resides in hypervariable gene families.

    18 January 2000

    • Ross Mayhew requests help in identifying this cone from Bohol Is. (collected at 35 fthms in December 1999). It has a uniform violet color, except just inside the lip, where the outer part is lighter. Note also the extremely regular spiral ridges, and the dashes of brown at the suture. Any cone-experts out there?

    • Added reference on the conopeptide content of Conus textile and Conus striatus using PCR amplification of alpha-conotoxin cDNA from venom duct cDNA, and rapid amplification of 3' cDNA ends (3'RACE) of 0-superfamily conotoxins:
      Lu Bai-Song, Yu fang, Shao Dong, Huang Pei-Tang, Juang Cui-Fen (1999)Conopeptides from Conus striatus and Conus textile by cDNA cloning. Peptides 20: 1139-1144.
      Abstract:Conopeptide content in Conus textile and Conus striatus venoms were examined by polymerase chain reaction amplification of alpha-conopeptide cDNA and rapid amplification of 3' cDNA ends of O-superfamily conopeptide cDNA. Two new alpha-conopeptide sequences and six new O-superfamily conopeptide sequences from C.textile, four new O-superfamily conopeptide sequences, and four previously biochemically characterized conopeptide sequences from C. striatus were identified. The results suggest that this cDNA method is rapid and requires less material for the study of conopeptides.

    9 January 2000

    • Our favourite mollusc, the cone shell Conus sp, has been noticed by Yolanda Fernandez, a reporter for The Alternative Medicine Report on WFLA-TV in Tampa, Florida. This article Sea Snail Venom, is primarily directed at those in pain who want to find out about ziconotide (conotoxin MVIIA) and pain prevention. A telephone contact number in Tampa, Florida, is provided for enquiries about clinical trials of ziconotide .
    • Two articles on pain, Living in Pain - Part 1 and Part 2, from PDX NORML, Portland, Oregon, retells the experience of Chris Ally from California, who had chronic pain and considered ziconotide but became anxious at the point of having a pump implanted and as a result, became ineligible for the ziconotide trial.
      These articles form part of a series reported and written with the cooperation of patients and their doctors, who were consulted throughout. To read the series online, log onto the San Francisco Chronicle Archives at www.sfgate.com or choose from the selected articles below:


    • Stephan Frings (from the Institut für Biologische Informationsverarbeitung, Jülich, Germany) has an interest in conotoxins and in teaching students about them.
      Take a look at some of his informative sites - and/or DOWNLOAD the scripts to your own computer (cono.exe 1.5Mb).
      Stephan's Homepage: "This website contains internet scripts written for students of biology at the University of Cologne, Germany. The scripts are mainly collections of figures used for teaching various topics of zoophysiology. Each figure can be viewed/loaded in a low-resolution (20-60 kbyte) and a high- resolution (>100 kbyte) version. At the moment, all accompanying texts are in German, but I will provide an english version in the future."

      Alternatively, you could translate the German into English (or to any other language of your choice !) with the Free Babel Fish add-in module for your web browser. Instructions are on the Babel Fish Tool page. Also see example "Five families of Conotoxins", below).

      View the article on Conotoxins in Boehringer magazine, FUTURA 12: 77-86, (1997). (Dont try translating the German abstract in this FUTURA article as the file is an image file only !).

      • Conotoxine - Muskelgifte der Kegelschnecken (November 1998)
          (http://www.zam.kfa-juelich.de/ibi/ibi-1/stefring/cono/conoin.htm)

        • Anatomie der Kegelschnecken Internal anatomy of the venom apparatus and radula sack of a cone snail.
          Also a collection of 6 cone shells.
        • Die Jagdstrategie von Conus purpurascens
          Conus purpurascens envenomating a fish (photos from Olivera's paper)
        • Das Gift der Kegelschnecken
          Two nice diagrams: One on milking cones. The other the HPLC trace of contoxins.

        • Fünf Familien von Conotoxinen

          - or as translated by Babel Fish Tool add-in to your web browser: When the screen appears just press "Translate" button.
          Five families of Conotoxins

          - or after a little bit of editing by BGL in Word of the above Babel Fish Tool translation.
          Five Conotoxin Families: Primary structures of the 5 families of conotoxins. High and low resolution.

        • Die Vergiftung der neuromuskulären Synapse
          - excellent diagram showing pre-synaptic and post-synaptic processes at nerve terminals and the interaction of conotoxins with the ion channels.
        • Physiologie der Schmerzwahrnehmung
          - excellent depiction of physiology and anatomy of neuronal pathways affected by conotoxins and interaction with morphine.
        • Conotoxine als Arznei
          - excellent diagram explaining the intrathecal injection of conotoxin for pain prevention

      • Literatur
        Selection of key conotoxin literature up to 1997

    8 January 2000

      Added reference to the first conotoxin paper for the year 2000 from Baldomero Olivera and his research team who examined six toxins for their ability to block different types of heterologously expressed sodium channels in a rat neuronal cell line (PC12) and in NGF-treated PC12 cells in which mRNAs for two neuronal voltage-activated sodium channels were induced. Their results provide support for the idea that two distinct sodium channel types (rBII/IIA and rPN1) appear sequentially during neuronal differentiation of PC12 cells:

    • Safo,P., Rosenbaum, T., Shcherbatko, A., Choi, D.-Y., Han, E., Toledo-Aral, J.J., Olivera, B.M., Brehm, P. and Mandel, G. (2000) Distinction among Neuronal Subtypes of Voltage-Activated Sodium Channels by µ-Conotoxin PIIIA
      The Journal of Neuroscience 20(1):76-80
      .
      Abstract: The functional properties of most sodium channels are too similar to permit identification of specific sodium channel types underlying macroscopic current. Such discrimination would be particularly advantageous in the nervous system in which different sodium channel family isoforms are coexpressed in the same cell. To test whether members of the µ-conotoxin family can discriminate among known neuronal sodium channel types, we examined six toxins for their ability to block different types of heterologously expressed sodium channels. PIIIA µ-conotoxin blocked rat brain type II/IIA (rBII/IIA) and skeletal muscle sodium current at concentrations that resulted in only slight inhibition of rat peripheral nerve (rPN1) sodium current. Recordings from variant lines of PC12 cells, which selectively express either rBII/IIA or rPN1 channel subtypes, verified that the differential block by PIIIA also applied to native sodium current. The sensitivity to block by PIIIA toxin was then used to discriminate between rBII/IIA and rPN1 sodium currents in NGF-treated PC12 cells in which both mRNAs are induced. During the first 24 hr of NGF-treatment, PN1 sodium channels accounted for over 90% of the sodium current. However, over the ensuing 48 hr period, a sharp rise in the proportion of rBII/IIA sodium current occurred, confirming the idea, based on previous mRNA measurements, that two distinct sodium channel types appear sequentially during neuronal differentiation of PC12 cells.
      Key words: PC12 cells; ion channel; sodium current; growth factor; CNS; PNS

    3 January 2000

    1 January 2000

      Happy New Year !

    25 December 1999

    21 December 1999

      Added reference to 'Toto' Olivera's thought-provoking review in Annals of the New York Academy of Sciences volume on "Molecular Strategies in Biological Evolution", Edited by Lynn Helena Caporale (Consultant, New York City), published as Proceedings of a New York Academy of Sciences conference, June 27-29, 1998.

      Outline This important reference work, the result of a conference co-chaired with Nobel laureate Werner Arber, addressed the molecular strategies by which lineages of organisms respond to challenges and opportunities in their environment. It explored the notion that organisms have evolved the ability to modulate the rate, location, and extent of genetic variation. Jumps in efficiency, made possible by development of novel, efficient evolutionary strategies, could fuel rapid, saltatory expansion of species into novel niches as each innovation evolves. An up-to-date assessment is provided on biochemical mechanisms available to modulate the rate of genetic change at specific sites within a genome, the induction in certain environments of enzymes with altered sequence-dependent recombination, mismatch repair and/or replication fidelity, and statistical evidence for non-random genetic events. This discussion of genomic strategies for evolution has profound implications for basic biology and evolutionary theory. The subjects explored are important ones in understanding inherited diseases, tumor progression, and the challenges posed by pathogenic organisms.
    • Olivera, B.M., Walker, C., Cartier, G.E., Hooper, D., Santos, A.D., Schoenfeld, R., Shetty, R., Watkins, M., Bandyopadyuay, P. and Hillyard, D.R.(1999) "Speciation of Cone Snails and Interspecific Hyperdivergence of their Venom Peptides: Potential Evolutionary Significance of Introns". Ann. N.Y. Acad. Sci. 870: 223-237.
      Abstract "All 500 species of cone snails (Conus) are venomous predators. From a biochemical / genetic perspective, differences among Conus species may be based on the 50-100 different peptides in the venom of each species. Venom is used for prey capture as well as for interactions with predators and competitors. The venom of every species has its own distinct complement of peptides. Some of the interspecific divergence observed in venom peptides can be explained by differential expression of venom peptide superfamilies in differnet species and of peptide superfamily branching in various Conus lineages into pharmacologic groups with different targeting specificity. However, the stiking interspecific divergence of peptide sequences is the dominant factor in the differentces observed between venoms. The small venom peptides (typically 10-35 amino acids in length) are processed from larger prepropeptide precursors (ca. 100 amino acids). If interspecific comparisons are made between homologous prepropeptides, the three different regions of a Conus peptide precursor (signal sequence, pro-region. mature peptide) are found to have diverged at remarkably different rates. Analysis of synonymous and nonsynonymous substitution rates for the different segments of a prepropeptide suggests that mutation frequency varies by over an order of magnitude across the segments, with the mature toxin region undergoing the highest rate. The three sections of the prepropeptide which exhibit apparently different mutation rates are separated by introns. This striking segment-specific rate of divergence of Conus prepropeptide suggests a role for introns in evolution: exons separated by introns have the potential to eveolve very different mutation rates. Plausible mechanisms that could underlie differing mutational frequency in the different exons of a gene are discussed .

    20 December 1999

      Added three references on the selectivity of alpha-Conotoxin PnIA/PnIB (from the molluscivorous cone snail Conus pennaceus) for the mammalian neuronal-type nicotinic acetylcholine receptor.

    • Broxton, N., Miranda, L., Gehrmann, J., Down, J., Alewood, P. and Livett, B.(2000) "Leu 10 of alpha-conotoxin PnIB confers potency for the neuronal nicotinic responses in bovine chromaffin cells". Europ. J. Pharmacol. 390: 229-236.
      Abstract "Two alpha-conotoxins PnIA and PnIB (previously reported as being "mollusc specific") which differ in only two amino acid residues (AN versus LS at residues 10 and 11, respectively), show markedly different inhibition of the neuronal nicotinic acetylcholine receptor response in bovine chromaffin cells, a mammalian preparation. Whereas alpha-conotoxin PnIB completely inhibits the nicotine-evoked catecholamine release at 10 uM, with IC50= 0.7 uM, alpha-conotoxin PnIA is some 30-40 times less potent. Two peptide analogues, [A10L]PnIA and [N11S]PnIA were synthesized to investigate the extent to which each residue contributes to activity. [A10L]PnIA (IC50=2.0uM) completely inhibits catecholamine release at 10 uM whereas [N11S]PnIA shows little inhibition. In contrast, none of the peptides inhibit muscle-type nicotinic responses in the rat hemi-diaphragm preparation. We conclude that the enhanced potency of alpha-conotoxin PnIB over alpha-conotoxin PnIA in the neuronal-type nicotinic response is principally determined by the larger, more hydrophobic leucine residue at position 10 in alpha-conotoxin PnIB."

    • Hogg, R.C., Miranda, L.P., Craik, D.J., Lewis, R.J., Alewood, P.F. and Adams, D.J.(1999) "Single amino acid substitutions in alpha-Conotoxin PnIA shift selectivity for subtypes of the mammalian neuronal nicotinic acetylcholine receptor" J.Biol.Chem. 274: 36559-36564.
      Abstract "The alpha-conotoxins, a class of nicotinic acetylcholine receptor (nAChR) antagonists, are emerging as important probes of the role played by different nAChR subtypes in cell function and communication. In this study, the native alpha-conotoxins PnIA and PnIB were found to cause concentration-dependent inhibition of the ACh-induced current in all rat parasympathetic neurons examined, with IC(50) values of 14 and 33 nM, and a maximal reduction in current amplitude of 87% and 71%, respectively. The modified alpha-conotoxin [N11S]PnIA reduced the ACh-induced current with an IC(50) value of 375 nM and a maximally effective concentration caused 91% block. [A10L]PnIA was the most potent inhibitor, reducing the ACh-induced current in ~80% of neurons, with an IC(50) value of 1.4 nM and 46% maximal block of the total current. The residual current was not inhibited further by alpha-bungarotoxin, but was further reduced by the alpha-conotoxins PnIA or PnIB, and by mecamylamine. (1)H NMR studies indicate that PnIA, PnIB, and the analogues, [A10L]PnIA and [N11S]PnIA, have identical backbone structures. We propose that positions 10 and 11 of PnIA and PnIB influence potency and determine selectivity among alpha7 and other nAChR subtypes, including alpha3beta2 and alpha3beta4. Four distinct components of the nicotinic ACh-induced current in mammalian parasympathetic neurons have been dissected with these conopeptides".

    • Luo, S., Nguyen, T.A., Cartier, G.E., Olivera, B.M., Yoshikami, D. and McIntosh, J.M. (1999) "Single-residue alteration in alpha-conotoxin PnIA switches its nAChR subtype selectivity". Biochemistry 38:14542-14548
      Abstract "alpha-Conotoxins are disulfide-rich peptides that are competitive antagonists of nicotinic acetylcholine receptors (nAChRs). Despite their small size, different alpha-conotoxins are able to discriminate among different subtypes of mammalian nAChRs. In this report, the activity of two peptides from the venom of Conus pennaceus, alpha-conotoxins PnIA and PnIB, are examined. Although the toxins differ in only two residues, PnIA preferentially blocks alpha3beta2 nAChRs, whereas PnIB prefers the alpha7 subtype. Point mutation chimeras of these alpha-conotoxins were synthesized and their activities assessed on Xenopus oocytes expressing specific nAChRs. Change of a single residue, Ala10 to Leu, in PnIA (to form PnIA [A10L]) converts the parent peptide from alpha3beta2-preferring to alpha7-preferring; furthermore, PnIA [A10L] blocks the alpha7 receptor with an IC(50) (12.6 nM) that is lower than that of either parent peptide. Kinetic analysis indicates that differences in affinity among the analogues are primarily due to differences in off-rate, with PnIA [A10L]'s interaction with alpha7 having the smallest off-rate (k(off) = 0.17 min(-)(1)). Thermodynamic analysis indicates that Leu10 enhances the peptide's interaction with alpha7, but not alpha3beta2, receptors, whereas Ser11 (in PnIA [N11S]) reduces its affinity for both alpha7 and alpha3beta2 nAChRs".

    17 December 1999

    • Added a reference from Bruce Furie's research group on the role of hydrophobic amino acids in precursor function in Conus textile.

      Bush, K.A., Stenflo, J., Roth, D.A., Czerwiec, E., Harrist, A., Begley, G.S., Furie, B.C. and Furie, B. (1999). Hydrophobic amino acids define the carboxylation recognition site in the precursor of the gamma- carboxyglutamic-acid-containing conotoxin epsilon-TxIX from the marine cone snail Conus textile. Biochemistry 38:14660-14666. To identify the amino acid sequence of the precursor of the Gla-containing peptide, epsilon-TxIX, from the venom of the marine snail Conus textile, the cDNA encoding this peptide was cloned from a Conus textile venom duct library. The cDNA of the precursor form of epsilon-TxIX encodes a 67 amino acid precursor peptide, including an N-terminal prepro-region, the mature peptide, and four residues posttranslationally cleaved from the C-terminus. To determine the role of the propeptide in gamma-carboxylation, peptides were designed and synthesized based on the propeptide sequence of the Gla-containing conotoxin epsilon-TxIX and used in assays with the vitamin K-dependent gamma-glutamyl carboxylase from C. textile venom ducts. The mature acarboxy peptide epsilon-TxIX was a high K(M) substrate for the gamma-carboxylase. Synthetic peptides based on the precursor epsilon-TxIX were low K(M) substrates (5 &mgr;M) if the peptides included at least 12 residues of propeptide sequence, from -12 to -1. Leucine-19, leucine-16, asparagine-13, leucine-12, leucine-8 and leucine-4 contribute to the interaction of the pro-conotoxin with carboxylase since their replacement by aspartic acid increased the K(M) of the substrate peptide. Although the Conus propeptide and the propeptides of the mammalian vitamin K-dependent proteins show no obvious sequence homology, synthetic peptides based upon the structure of pro-epsilon-TxIX were intermediate K(M) substrates for the bovine carboxylase. The propeptide of epsilon-TxIX contains significant alpha-helix, as estimated by measurement of the circular dichroism spectra, but the region of the propeptide that plays the dominant role in directing carboxylation does not contain evidence of helical structure. These results indicate that the gamma-carboxylation recognition site is defined by hydrophobic residues in the propeptide of this conotoxin precursor.

    12 December 1999

    • Updated the list of Conotoxin Patents to include the following:
      WO09954350A1 "Novel Conotoxin Peptides" by Roger Drinkwater, Richard Lewis, Paul Alewood and Kathy Nielsen, Centre for Drug Design and Development, University of Queensland, Australia. [An isolated, synthetic or recombinant omega-conotoxin peptide in which the fourth loop between cysteine residues 5 and 6 comprises SEQ ID NO: 1 (SEQ ID NO: 1 comprises the amino acid sequence SGTVGR ) or such a sequence which has undergone one or more amino acid substitutions or side chain modifications, and uses therefor. The invention relates to novel peptides of the omega conotoxin class and their use as pharmacological tools in any indication where blockage of N-type calcium channels may be of benefit, for example in the reduction of neuronal damage following ischemia, production of analgesia, or enhancement of opiate analgesia, in the treatment of schizophrenia, stimulant induced psychoses, hypertension, inflammation and diseases which cause bronchoconstriction and in the inbibition of progression of neuropathic pain. Information, IBM Patent Searcher and WIPO PCT Publications]

    5 December 1999

    • Added two references on cone shell toxins (conotoxins) and their interactions with potassium ion channels.
      Heinrich Terlau, Anna Boccaccio, Baldomero M. Olivera, and Franco Conti (1999) The Block of Shaker K+ Channels by k-Conotoxin PVIIA Is State Dependent. J. Gen. Physiol. 114: 125-140.
      kappa-conotoxin PVIIA is the first conotoxin known to interact with voltage- gated potassium channels by inhibiting Shaker-mediated currents. We studied the mechanism of inhibition and concluded that PVIIA blocks the ion pore with a 1:1 stoichiometry and that binding to open or closed channels is very different. Open-channel properties are revealed by relaxations of partial block during step depolarizations, whereas double-pulse protocols characterize the slower reequilibration of closed-channel binding. In 2.5 mM-[K+]o, the IC50 rises from a tonic value of approximately 50 to approximately 200 nM during openings at 0 mV, and it increases e-fold for about every 40-mV increase in voltage. The change involves mainly the voltage dependence and a 20-fold increase at 0 mV of the rate of PVIIA dissociation, but also a fivefold increase of the association rate. PVIIA binding to Shaker Delta6-46 channels lacking N-type inactivation or to wild phenotypes appears similar, but inactivation partially protects the latter from open-channel unblock. Raising [K+]o to 115 mM has little effect on open-channel binding, but increases almost 10-fold the tonic IC50 of PVIIA due to a decrease by the same factor of the toxin rate of association to closed channels. In analogy with charybdotoxin block, we attribute the acceleration of PVIIA dissociation from open channels to the voltage-dependent occupancy by K+ ions of a site at the outer end of the conducting pore. We also argue that the occupancy of this site by external cations antagonizes on binding to closed channels, whereas the apparent competition disappears in open channels if the competing cation can move along the pore. It is concluded that PVIIA can also be a valuable tool for probing the state of ion permeation inside the pore.

      Esperanza García, Martin Scanlon, and David Naranjo (1999) A Marine Snail Neurotoxin Shares with Scorpion Toxins a Convergent Mechanism of Blockade on the Pore of Voltage-gated K Channels.J. Gen. Physiol. 114: 141-158.
      kappa-Conotoxin-PVIIA (kappa-PVIIA) belongs to a family of peptides derived from a hunting marine snail that targets to a wide variety of ion channels and receptors. kappa-PVIIA is a small, structurally constrained, 27-residue peptide that inhibits voltage-gated K channels. Three disulfide bonds shape a characteristic four-loop folding. The spatial localization of positively charged residues in kappa-PVIIA exhibits strong structural mimicry to that of charybdotoxin, a scorpion toxin that occludes the pore of K channels. We studied the mechanism by which this peptide inhibits Shaker K channels expressed in Xenopus oocytes with the N-type inactivation removed. Chronically applied to whole oocytes or outside-out patches, kappa-PVIIA inhibition appears as a voltage-dependent relaxation in response to the depolarizing pulse used to activate the channels. At any applied voltage, the relaxation rate depended linearly on the toxin concentration, indicating a bimolecular stoichiometry. Time constants and voltage dependence of the current relaxation produced by chronic applications agreed with that of rapid applications to open channels. Effective valence of the voltage dependence, zdelta, is approximately 0.55 and resides primarily in the rate of dissociation from the channel, while the association rate is voltage independent with a magnitude of 10(7)-10(8) M-1 s-1, consistent with diffusion-limited binding. Compatible with a purely competitive interaction for a site in the external vestibule, tetraethylammonium, a well-known K-pore blocker, reduced kappa-PVIIA's association rate only. Removal of internal K+ reduced, but did not eliminate, the effective valence of the toxin dissociation rate to a value <0.3. This trans-pore effect suggests that: (a) as in the alpha-KTx, a positively charged side chain, possibly a Lys, interacts electrostatically with ions residing inside the Shaker pore, and (b) a part of the toxin occupies an externally accessible K+ binding site, decreasing the degree of pore occupancy by permeant ions. We conclude that, although evolutionarily distant to scorpion toxins, kappa-PVIIA shares with them a remarkably similar mechanism of inhibition of K channels.

    • Linked to Gary Rosenberg's marvelous "Conchatenations" column on the question Mollusk or Mollusc ? - [from the September 1999 issue of American Conchologist].

    Continued in What's new in 1999


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