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ω-Conotoxins: calcium channel inhibitors from Conus venoms
S00
Tenth European Sympaium
(attenuated total reflection) gave information about the oriaatation of DT in the lipid matrQ. On the basis of the expe
Study ojstrvctroe-actldty rc/atfonahip ofjarcfculln by chnntcd nw~~COtioh ojaglniree, lyaihe mtd cmboxylate groups. C. L`mrv~~f,' F. DAIAe,' A. Exostxôsrr and E. R~ar.~ox' ('Imtituto de Invetigaciona Hiol6gicas Ckmeate Eatable, Divisi6n Neuroquimica, Avenida Italie 3318, 11600 Montevideo, Uruguay; fiepartmeat of Immunology, Biomedical Cmtro, Box 582, 75123 Uppsala, Sweden; and ~Department of Hiorbemietry, Biomedical Centre, Box 576, 75123 Uppeala, Sweden). Fesctan.uva (ZerveBaaskj~ et d. 1991) from the venom of the wake D~adroaapta arrgruticeps (green mamba) are 61 amino acid residue peptide structurally homologous to short-chain a-neurotoxins. They are potent noncompetitive inhibitors of many aoetylcholineterase (~ s 0.13 nM for electric eel enzyme) and bind to a peripheral aaionic site . Chemical modifications of fasciculin 2 were carried out in order to study the role of the flue amino groups, six argininea and six carboxyls. Amino groups were acetylated with rl-I-acetic anhydride and the frn monoaoetyl derivatives were purified by HPLC cation-exchange chromstographY. When the 1abe1 was on Lye-2S or Lys-32, the remaining activity was about 305.. The other derivatives were moro activa Arginine residue were modified wiW 1,2-cyclohexanedione. Of the flue mono-substituted derivative, twro had the same activity as the native toxin, two had about 2Sy. remaining activity and one ler than IS'/.. Carboxylic groups were modified by coupling them to norleuciae methylester with a carbodümide. All mono-derivatives had about the same activity as the native toxin. These results i~te that many cationic groups in the faecicailin molecule are involved in the interaction with acetylcholinesterase. ~rvsEt~f, C. et d. (1991) Fasciculins, anticholineterase toxins from mamba venoma : biochemistry and pharmacology. In: Snake Toxinr, Internationd Facydopedio of Pharmacology road Thaoptutks, pp . 301-321 (Harvey, A. L., Ed.). New York: Pergamon Press. This work was supported by IPICS (LJppsala University), IFS (Stockholm), The Swedish Medical Research Council and The Swedish Natural Seisms Reearch Couadl. arCarotoxinu : cdc8wt demsnel tnhibftors fron Conua vasams. L. J. Cauz,'a C. ZAFARA~A, ' V. D. Mowe,~ B. M. Otav®u' and D. R Hn r vNrn' ('Marine Science Institute and rInatitute of Chemistry University of the Phr7ippine, Dilimsa, QC 1101, Philippine ; and'Department of )äology, and pathology, University of Utah, Salt Lake sty, UT 84112, U.SA.). orCotro'roxnv ßVIA from Cotars geographtcs is now wily used as a pharmacological tool for identifying the N-subtype of Ca charnels, which controls neurotransmitter release is certain cell type of the mammalian CNS. The peptide induces ehalting when injected intracraniallY in mice. In fish, frogs, and chicks, the toxin cause P~Y~ bY blocking pteaynaptic Ca channels of the neuromuscular junction . Reoeatly, two Ca channel inhibitors, aroonotoxins SVIA a~ SVIB have been isolated from C. striatue (Reim.o et d., 1992). Although SVIA is a potent paralytic toxin in lower vertebrate specie, it ie much ler effective than GVIA in mammals with respect to the shaker activity. On the other hand, of the 11 erconotoxins so far purified from COtnLS venom, the shaker peptide SV1B it the only variant lethal to mice oa i.c . injection. Binding competition experiments with rat brain synaptosomal membrane indicate that the high affinity site for SVIB ie distinct from the high a®nity site for ßVIA of C. geographw and/or MV17A of C. magus. The application of molecular biological technique has revealed another variant of arconotoxin not yet purified from Come venom (Hnl.v~an et d., 1992). This peptide, arconotoxin MVIIC, was identified from a C. magna cDNA library. Chemical synthesis of the predicted translation product gave a peptide which û lethal to mice without the accompanying abaking syndrome. The ar GVIA-tssistant Ca channels inhibited by arMVIIC include those that mediate depolarization-induced "Ca uptake in rat synaptosome pmparations, 'P' currents in oerebelhu Purkinje ceW, sad a wbset of m-ßVIAroristant currents in CAl hippocampal PYramidal ceW (Ha.t.Yerin et d., 1992). The only absolutely conserved residue in the thirteen known variants of aroonotoxins aro the six Cys residue and Glys; the residue before the last cyeteine can either be Lys or Arg. Comparison of natural sequence as well as an extemive analysis of synthetic analogs (J . Rtiucrurmawrr, ß. Ma.r~nctt and L. Nenasor, unpublished results) indicate that the fast three residues of loop 2 may be important for determining Ca subtype specificity. Reim.o et d. (1992) Biodianistry, in pr+es. Ha.r .vean et d. (1992) Neivoti 9, 69-77. Supported by NIIi grant GM22737 to HMO and by grants from the Marine Scimoe Inst ., U.P. Diliman, Philippines; and the International Foundation for Science, Stockholm, Sweden to LJC. Cobra rtnom plwsphOllpasc A7 structwt and nitdiossLnri . E. A. D~ri~ns (Department of Chemistry, 0601,
University of California at San Diems, La Jolle, CA 92093-0601, U.S.A .) .
Tenth European Sympaium
(attenuated total reflection) gave information about the oriaatation of DT in the lipid matrQ. On the basis of the expe
Study ojstrvctroe-actldty rc/atfonahip ofjarcfculln by chnntcd nw~~COtioh ojaglniree, lyaihe mtd cmboxylate groups. C. L`mrv~~f,' F. DAIAe,' A. Exostxôsrr and E. R~ar.~ox' ('Imtituto de Invetigaciona Hiol6gicas Ckmeate Eatable, Divisi6n Neuroquimica, Avenida Italie 3318, 11600 Montevideo, Uruguay; fiepartmeat of Immunology, Biomedical Cmtro, Box 582, 75123 Uppsala, Sweden; and ~Department of Hiorbemietry, Biomedical Centre, Box 576, 75123 Uppeala, Sweden). Fesctan.uva (ZerveBaaskj~ et d. 1991) from the venom of the wake D~adroaapta arrgruticeps (green mamba) are 61 amino acid residue peptide structurally homologous to short-chain a-neurotoxins. They are potent noncompetitive inhibitors of many aoetylcholineterase (~ s 0.13 nM for electric eel enzyme) and bind to a peripheral aaionic site . Chemical modifications of fasciculin 2 were carried out in order to study the role of the flue amino groups, six argininea and six carboxyls. Amino groups were acetylated with rl-I-acetic anhydride and the frn monoaoetyl derivatives were purified by HPLC cation-exchange chromstographY. When the 1abe1 was on Lye-2S or Lys-32, the remaining activity was about 305.. The other derivatives were moro activa Arginine residue were modified wiW 1,2-cyclohexanedione. Of the flue mono-substituted derivative, twro had the same activity as the native toxin, two had about 2Sy. remaining activity and one ler than IS'/.. Carboxylic groups were modified by coupling them to norleuciae methylester with a carbodümide. All mono-derivatives had about the same activity as the native toxin. These results i~te that many cationic groups in the faecicailin molecule are involved in the interaction with acetylcholinesterase. ~rvsEt~f, C. et d. (1991) Fasciculins, anticholineterase toxins from mamba venoma : biochemistry and pharmacology. In: Snake Toxinr, Internationd Facydopedio of Pharmacology road Thaoptutks, pp . 301-321 (Harvey, A. L., Ed.). New York: Pergamon Press. This work was supported by IPICS (LJppsala University), IFS (Stockholm), The Swedish Medical Research Council and The Swedish Natural Seisms Reearch Couadl. arCarotoxinu : cdc8wt demsnel tnhibftors fron Conua vasams. L. J. Cauz,'a C. ZAFARA~A, ' V. D. Mowe,~ B. M. Otav®u' and D. R Hn r vNrn' ('Marine Science Institute and rInatitute of Chemistry University of the Phr7ippine, Dilimsa, QC 1101, Philippine ; and'Department of )äology, and pathology, University of Utah, Salt Lake sty, UT 84112, U.SA.). orCotro'roxnv ßVIA from Cotars geographtcs is now wily used as a pharmacological tool for identifying the N-subtype of Ca charnels, which controls neurotransmitter release is certain cell type of the mammalian CNS. The peptide induces ehalting when injected intracraniallY in mice. In fish, frogs, and chicks, the toxin cause P~Y~ bY blocking pteaynaptic Ca channels of the neuromuscular junction . Reoeatly, two Ca channel inhibitors, aroonotoxins SVIA a~ SVIB have been isolated from C. striatue (Reim.o et d., 1992). Although SVIA is a potent paralytic toxin in lower vertebrate specie, it ie much ler effective than GVIA in mammals with respect to the shaker activity. On the other hand, of the 11 erconotoxins so far purified from COtnLS venom, the shaker peptide SV1B it the only variant lethal to mice oa i.c . injection. Binding competition experiments with rat brain synaptosomal membrane indicate that the high affinity site for SVIB ie distinct from the high a®nity site for ßVIA of C. geographw and/or MV17A of C. magus. The application of molecular biological technique has revealed another variant of arconotoxin not yet purified from Come venom (Hnl.v~an et d., 1992). This peptide, arconotoxin MVIIC, was identified from a C. magna cDNA library. Chemical synthesis of the predicted translation product gave a peptide which û lethal to mice without the accompanying abaking syndrome. The ar GVIA-tssistant Ca channels inhibited by arMVIIC include those that mediate depolarization-induced "Ca uptake in rat synaptosome pmparations, 'P' currents in oerebelhu Purkinje ceW, sad a wbset of m-ßVIAroristant currents in CAl hippocampal PYramidal ceW (Ha.t.Yerin et d., 1992). The only absolutely conserved residue in the thirteen known variants of aroonotoxins aro the six Cys residue and Glys; the residue before the last cyeteine can either be Lys or Arg. Comparison of natural sequence as well as an extemive analysis of synthetic analogs (J . Rtiucrurmawrr, ß. Ma.r~nctt and L. Nenasor, unpublished results) indicate that the fast three residues of loop 2 may be important for determining Ca subtype specificity. Reim.o et d. (1992) Biodianistry, in pr+es. Ha.r .vean et d. (1992) Neivoti 9, 69-77. Supported by NIIi grant GM22737 to HMO and by grants from the Marine Scimoe Inst ., U.P. Diliman, Philippines; and the International Foundation for Science, Stockholm, Sweden to LJC. Cobra rtnom plwsphOllpasc A7 structwt and nitdiossLnri . E. A. D~ri~ns (Department of Chemistry, 0601,
University of California at San Diems, La Jolle, CA 92093-0601, U.S.A .) .