Department of Surgery, Yijishan Hospital, Wannan Medical College, Wuhu, Anhui 241001, China.
Chin Med J (Engl). 2009 Sep 20;122(18):2169-73.
Snake venom contains a number of components with different pharmacological and biological activities, especially in cancer therapy, and has increasingly become a research focus. This study was designed to isolate and purify a novel anti-clotting protein component from the venom of Agkistrodon acutus, and to explore its physico-chemical properties and biological activity.
The venom of Agkistrodon was isolated and purified by ion-exchange chromatography on diethylaminoethyl (DEAE)-Sepharose Fast Flow, molecular sieve filtration through Sephadex G75, SP-Sepharose Fast Flow and molecular sieve filtration through Sephadex G50. We detected the activated partial thromboplastin time (APTT) of the eluant to select the anti-clotting protein component of interest. The molecular weight was determined by sodium dodecyl sulfate-polyacrylamid gel electrphoresis (SDS-PAGE) and liquid chromatography. Its protein content was detected by bicinchoninic acid (BCA).
SDS-PAGE vertical gel electrophoresis showed that the anticoagulant factor is a tripolymer composed of three proteins whose molecular weights are 25 KDa, 30 KDa and 50 KDa. The factor contains about 65% percent protein.
A novel anti-clotting protein component was purified by ion-exchange chromatography and molecular sieve filtration from the venom of Agkistrodon acutus and was found to be composed of three kinds of proteins.
蛇毒含有多种具有不同药理和生物学活性的成分，尤其在癌症治疗方面，越来越成为研究热点。本研究旨在从尖吻蝮蛇毒中分离纯化一种新型抗凝血蛋白成分，并对其理化性质和生物学活性进行研究。
采用离子交换层析 DEAE- Sepharose Fast Flow 分离、分子筛层析 Sephadex G75 进一步纯化尖吻蝮蛇毒，SP-Sepharose Fast Flow 离子交换层析和分子筛层析 Sephadex G50 分离纯化抗凝血蛋白成分，通过检测洗脱液对活化部分凝血活酶时间（APTT）的影响，筛选目的抗凝血蛋白成分。采用十二烷基硫酸钠-聚丙烯酰胺凝胶电泳（SDS-PAGE）和液相色谱法测定其相对分子质量，考马斯亮蓝法（BCA）测定蛋白含量。
SDS-PAGE 垂直电泳显示，抗凝因子是由分子量分别为 25 kDa、30 kDa 和 50 kDa 的三种蛋白组成的三聚体，该因子约含有 65%的蛋白质。
采用离子交换层析和分子筛层析从尖吻蝮蛇毒中分离纯化得到一种新型抗凝血蛋白成分，该成分由三种蛋白组成。
