广西圆斑蝰蛇毒凝血因子X激活剂的分离纯化及其止血作用的研究(1)
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【摘要】 目的 从广西圆斑蝰蛇毒中分离纯化凝血因子X激活剂(factor X activator of Russell’s viper venom,RVV-X)并研究其止血作用。方法 应用阴离子交换色谱和凝胶过滤从广西圆斑蝰蛇毒中分离纯化RVV-X,以复钙化时间进行活性鉴定,以SDS-PAGE测定纯度和分子量。观察RVV-X对家兔肝脏创伤出血模型出血时间及出血量,家兔和肝素化小鼠凝血时间的影响。结果 广西圆斑蝰粗毒经阴离子交换色谱获得六个蛋白峰,将RVV-X所在第V峰用凝胶过滤获得的组分A为RVV-X纯品,其凝血活性具有钙依赖性。SDS-PAGE非还原性条件下呈单一蛋白条带,分子量约为97.8kDa,还原性条件下呈二蛋白条带,为一条重链(约76.3kDa)和一条轻链(约21.5kDa)。RVV-X可显著减少家兔肝脏切口的失血量,缩短切口出血时间;能显著缩短家兔及肝素化小鼠凝血时间。结论 用层析方法从广西圆斑蝰蛇毒中分离得到的凝血因子X激活剂具有较强的止血活性。
【关键词】 凝血因子X激活剂;离子交换;凝胶过滤;止血作用;圆斑蝰
Purification and Haemostatic Effect of Factor X activator from venom of Vipera russellii in Guangxi
XIE Yun,XU Yun-Lu.
Laboratory of Snake Venom Research,Fujian Medical University,Fuzhou350004,China
【Abstract】 Objective To isolate and purify the coagulation factor X activator from venom of Vipera russellii in Guangxi and to investigate its hemostasis effects.Methods RVV-X was purified from Russell’s viper venom by ion-exchange chromatography and by gel filtration. Homogenicity and molecular weight of it were determined in SDS-PAGE.Then we observed effects of intravenous RVV-X on model of hepatic trauma rabbit , on blood coagulation time in rabbit and heparinized mice. Results RVV-X was isolated from the venom of Vipera russellii.It could shorten the coagulation time in the presence of Ca.2+ ions.It consisted of a single polypeptide chain, with a mol. wt of 97,800 in nonreducing conditions and composed of one heavy chain (76,300Da) and one light chain(21,500Da) in reducing conditions.RVV-X could obviously decrease blood loss of rabbits liver wound and shorten bleeding time.It could also shorten the coagulation time of rabbit and heparinized mice. Conclusion RVV-X extracted from the venom of Vipera Russellii in Guangxi possesses considerable hemostatic activities.
【Key words】 Factor X activator;Ion-exchange chromatography;Gel filtration;Hemostasis effect;Vipera russellii
作者单位:350004福建医科大学药学院蛇毒研究所
蛇毒中含有多种可以影响哺乳动物凝血系统的酶类,其中在蝰亚科、蝮亚科、眼镜蛇科等蛇毒中存在着一类能激活凝血因子X的促凝组分-FX激活剂。国外对FX激活剂的研究主要集中在分离纯化、分子结构、凝血机制等方面[1~4],国内也报道了其在实验鼠体内的具有促凝血作用[5],但至今尚无FX激活剂作为止血药被开发上市 ......
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