Isolation and cloning of a metalloproteinase from king cobra snake venom
Isolation and cloning of a metalloproteinase from king cobra snake venom
复制标题
眼镜王蛇毒液中金属蛋白酶的分离和克隆
DOI：
10.1016/j.toxicon.2007.01.003
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发表时间：
2007-06-01
期刊：
影响因子：
2.8
通讯作者：
Jin, Yang
中科院分区：
文献类型：
--
作者：
Guo, Xiao-Xi;Zeng, Lin;Jin, Yang
采用凝胶过滤、离子交换和肝素亲和层析相结合的方法，从眼镜王蛇毒中分离得到一种50 kDa的纤溶蛋白酶ohagin。Ohagin特异性地降解人纤维蛋白原的α链，并且蛋白水解活性被EDTA完全消除，但不被PMSF消除，这表明它是一种金属蛋白酶。对ADP、TMVA和stejnulxin诱导的血小板聚集有剂量依赖性的抑制作用。通过cDNA克隆推导出ohagin的全序列，并通过蛋白质测序和肽质量指纹图谱进行了验证。ohagin全长cDNA序列编码611个氨基酸的开放阅读框，包括信号肽、前蛋白和成熟蛋白，成熟蛋白包括金属蛋白酶、去整合素样结构域和富含半胱氨酸的结构域，属于P-Ⅲ类金属蛋白酶。此外，本研究还从眼镜蛇、银环蛇和金环蛇的毒腺中克隆了P-Ⅲ类金属环蛋白酶。序列分析和系统发育分析表明，眼镜蛇蛇毒金属蛋白酶形成一个新的亚类P-III SVMP。(C)2007爱思唯尔有限公司版权所有。
A 50 kDa fibrinogenolytic protease, ohagin, from the venom of Ophiophagus hannah was isolated by a combination of gel filtration, ion-exchange and heparin affinity chromatography. Ohagin specifically degraded the alpha-chain of human fibrinogen and the proteolytic activity was completely abolished by EDTA, but not by PMSF, suggesting it is a metalloproteinase. It dose-dependently inhibited platelet aggregation induced by ADP, TMVA and stejnulxin. The full sequence of ohagin was deduced by cDNA cloning and confirmed by protein sequencing and peptide mass fingerprinting. The full-length cDNA sequence of ohagin encodes an open reading frame of 611 amino acids that includes signal peptide, proprotein and mature protein comprising metalloproteinase, disintegrin-like and cysteine-rich domains, suggesting it belongs to P-III class metalloproteinase. In addition, P-III class metal lopro tei nases from the venom glands of Naja atra, Bungarus multicinctus and Bungarus fasciatus were also cloned in this study. Sequence analysis and phylogenetic analysis indicated that metalloproteinases from elapid snake venoms form a new subgroup of P-III SVMPs. (C) 2007 Elsevier Ltd. All rights reserved.