The complete covalent structure of a cardiotoxin from the venom of Naja nigricollis (African black-necked spitting cobra).
The complete covalent structure of a small, basic protein with cardiotoxic activity is described. This has been isolated from the venom of Naja nigricollis by gel filtration on Sephadex G-75 and gradient ion exchange chromatography on Bio-Rex 70. The cardiotoxin, molecular weight 6806 from amino acid composition, consists of 60 amino acids, cross-linked by four disulfide bridges, connecting 3-21, 14-38, 42-53, and 54-59. The protein contains one residue of tryptophan, phenylalanine, and glutamic acid, two residues of arginine and tyrosine, four residues of methionine, and nine residues of lysine. Histidine is absent. The chymotryptic peptides of the oxidized and S-carboxymethylated protein were isolated by gel filtration on Sephadex G-25 and zone electrophoresis on a cellulose column. The sequence was determined by Edman degradation, using the (manual) direct phenylthiohydantoin method and with the use of carboxypeptidase A. Disulfide pairing was determined on thermolysin cleaved peptides from the native protein. The sequence is shown to be homologous to other cardiotoxins and a lytic factor from snake venoms and also shows homology, both in sequence and disulfide pairing to neurotoxins. A partial reduction experiment in the absence of denaturing agent using 14-C-labeled iodoacetic acid as S-carboxymethylating agent shows that disulfide bonds 14-38 and 42-53 were reduced fastest followed marginally by 54-59, and then bond 3-21.
D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D000818:Animals; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D002918:Chymotrypsin / Q000494:pharmacology; D003545:Cysteine / Q000378:metabolism; D004220:Disulfides / Q000032:analysis; D004586:Electrophoresis; D006321:Heart / Q000187:drug effects; D007526:Isoelectric Point; D010446:Peptide Fragments / Q000032:analysis; D012910:Snake Venoms / Q000032:analysis* / Q000494:pharmacology; D013820:Thermolysin / Q000494:pharmacology; D014357:Trypsin / Q000494:pharmacology; D014688:Venoms / Q000032:analysis*
D000596:Amino Acids; D004220:Disulfides; D010446:Peptide Fragments; D012910:Snake Venoms; D014688:Venoms; D002918:Chymotrypsin; D014357:Trypsin; D013820:Thermolysin; D003545:Cysteine