title: Characterization of a thrombin-like serine protease, Kangshuanmei, isolated from the venom of a Chinese snake, Agkistrodon halys brevicaudus stejneger.
abstract: An enzyme, referred to as Kangshuanmei, was isolated from the venom of the Chinese snake Agkistrodon halys brevicaudus stejneger by gel filtration chromatography followed by affinity chromatography. Kangshuanmei is composed of a single polypeptide chain with a molecular weight of approximately 34,000, estimated by SDS-PAGE. The enzyme hydrolyzed both benzoyl-arginine ethyl ester and H-D-Phe-Pip-Arg-p-nitroanilide, specific substrates for thrombin. The protease activity of Kangshuanmei was inhibited by 4-(2-aminoethyl)-benzensulfonyl fluoride, but was not affected by EDTA. The enzyme acted on human fibrinogen to form a fibrin clot and released three fragments. These fragments were shown to be fibrinopeptide A, fibrinopeptide B, and the Bbeta1-42 peptide of fibrinogen, respectively. These results indicate that Kangshuanmei is a thrombin-like serine protease with coagulant activity. However, the enzyme did not induce activation of blood coagulation factor XIII, unlike thrombin. Moreover, antithrombin-III, the specific thrombin inhibitor in plasma, had no inhibitory effect on the thrombin-like amidolytic activity of Kangshuanmei. The N-terminal amino acid sequence of the enzyme up to 50 residues was determined by a peptide sequencer. The N-terminal sequence of Kangshuanmei was highly homologous to most thrombin-like serine proteases from the venom of the snakes of the crotalidae family.
authors.affiliation: Department of Physiology, School of Medicine, Aichi Medical University, Nagakute, Japan.
authors.forename: S
authors.initials: S
authors.lastname: Zhang
authors.forename: B
authors.initials: B
authors.lastname: Ma
authors.forename: J
authors.initials: J
authors.lastname: Sakai
authors.forename: H
authors.initials: H
authors.lastname: Shiono
authors.forename: T
authors.initials: T
authors.lastname: Matsui
authors.forename: I
authors.initials: I
authors.lastname: Sugie
authors.forename: T
authors.initials: T
authors.lastname: Okada
mesh_terms: D017836:Agkistrodon; D000595:Amino Acid Sequence; D000818:Animals; D001777:Blood Coagulation / Q000187:drug effects*; D003435:Crotalid Venoms / Q000201:enzymology*; D004591:Electrophoresis, Polyacrylamide Gel; D005340:Fibrinogen / Q000378:metabolism; D006490:Hemostatics / Q000737:chemistry* / Q000302:isolation & purification* / Q000494:pharmacology; D006801:Humans; D008969:Molecular Sequence Data; D012697:Serine Endopeptidases / Q000737:chemistry* / Q000302:isolation & purification*; D013917:Thrombin / Q000737:chemistry* / Q000302:isolation & purification* / Q000494:pharmacology
chemical_list: D003435:Crotalid Venoms; D006490:Hemostatics; D005340:Fibrinogen; D012697:Serine Endopeptidases; C441966:kangshuanmei; D013917:Thrombin