Purification and amino acid sequence of halystase from snake venom of Agkistrodon halys blomhoffii, a serine protease that cleaves specifically fibrinogen and kininogen

Taei Matsui, Yoshihiko Sakurai, Yoshihiro Fujimura, Izumi Hayashi, Sachiko Oh-Ishi, Masami Suzuki, Jiharu Hamako, Yoshinobu Yamamoto, Junko Yamazaki, Michiko Kinoshita, Koiti Titani

研究成果: Article査読

40 被引用数 (Scopus)

抄録

We have isolated a serine protease, halystase, from Agkistrodon halys blomhoffii venom by chromatography on DEAE-Sepharose, heparin-Sepharose and Q-Sepharose columns, and have determined the complete amino acid sequence by Edman degradation and by mass spectral analysis of peptides generated by enzymatic and chemical cleavage. The 238-residue sequence of halystase, containing N-linked carbohydrates (about 13%) at two sites showed significant similarity to other thrombin-like snake venom serine proteases (66-72%), mammalian tissue kallikrein (42%) and thrombin (26%). Halys as contained the tentative catalytic triad of His43, Asp88 and Ser184 common to all serine proteases and Asp178 in the primary substrate-binding site. Although halystase contained an RGD sequence at residues 181-183, it did not inhibit platelet aggregation induced by ADP or collagen. It hydrolyzed most efficiently a tissue-kallikrein substrate, prolylphenylalanylarginyl-4- methyl-coumaryl-7-amide, and released bradykinin from bovine kininogen. Halystase did not coagulate human plasma, but it cleaved the fibrinogen B β chain at the carboxyl side of Arg42 and cleaved slowly the fibrogen A α chain. Fibrinogen thus treated gradually became insensitive to thrombin. The proteolytic activity was inhibited with diisopropyl fluorophosphate, phenylmethylsulfonyl fluoride or leupeptin. These results indicate that halystase is a serine protease structurally similar to coagulating thrombin- like snake venom proteases, but it specifically cleaves fibrinogen at sites different from thrombin without inducing fibrin clotting, and hydrolyzes kininogen to produce bradykinin, resulting in the reduction of blood pressure.

本文言語English
ページ(範囲)569-575
ページ数7
ジャーナルEuropean Journal of Biochemistry
252
3
DOI
出版ステータスPublished - 15-03-1998

All Science Journal Classification (ASJC) codes

  • 生化学

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