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|Title:||PURIFICATION, CHARACTERIZATION, and AMINO-ACID-SEQUENCE of A SERINE PROTEINASE, PA-BJ, WITH PLATELET-AGGREGATING ACTIVITY FROM the VENOM of BOTHROPS-JARARACA|
MAX PLANCK INST BIOCHEM
Universidade Federal de São Paulo (UNIFESP)
|Publisher:||Amer Chemical Soc|
|Citation:||Biochemistry. Washington: Amer Chemical Soc, v. 34, n. 21, p. 7186-7193, 1995.|
|Abstract:||A platelet-aggregating enzyme, PA-BJ, was isolated from the venom of the snake Bothrops jararaca. PA-BJ in a concentration of 3.2 x 10(-8) M promoted 95% platelet aggregation in platelet-rich plasma. SDS-polyacrylamide gel electrophoresis under reducing conditions showed a single protein band with an M(r) of 30 000. PA-BJ catalyzed the hydrolysis of several p-nitroanilide peptide substrates containing Arg or Lys at the scissile bond; among these the most sensitive were the thrombin substrates D-Phe-Pip-Arg-pNA and Tos-Gly-Pro-Arg-pNA. Both the platelet-aggregating and amidolytic activities of PA-BJ were abolished by reaction with phenylmethanesulfonyl fluoride. Several benzamidine derivatives, which are competitive inhibitors of trypsin-like serine proteinases, also inhibited the amidolytic activity of PA-BJ. Among the compounds tested, the thrombin inhibitor NAPAP [((alpha)-[(2-naphthylsulfonyl)- glycyl]-4-amidinophenylalanine piperidide] showed the strongest inhibitor activity on PA-BJ. the complete amino acid sequence of PA-BJ, which, to the best of our knowledge, is the first of a platelet-aggregating enzyme from snake venom, was deduced from the N-terminal sequencing of overlapping fragments cleaved from the reduced and S-pyridylethylated protein by chemical and enzymatic methods. PA-BJ is composed of 232 amino acid residues and contains one N- and one O-glycosidically linked carbohydrate moiety at residues Asn(20) and Ser(23). Sequence comparison to other venom serine proteinases revealed significant homology, mainly in regions around the catalytic triad and conserved cysteine residues.|
|Appears in Collections:||Em verificação - Geral|
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