色谱

色谱

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两步柱色谱法分离纯化重组肿瘤血管生长抑制因子Kringle 5

贾信贵 边六交   

  1. National Engineering Research Center for Miniaturized Detection System, College of Life Science, Northwest University, Xi’an 710069, China
  • 收稿日期:2006-10-15 修回日期:2006-11-21 出版日期:2007-05-30 发布日期:1985-09-25
  • 通讯作者: 边六交

Two-Step Chromatographic Method for the Separation and Purification of Recombinant Angiogenesis Inhibitor Kringle 5

JIA Xingui, BIAN Liujiao   

  1. National Engineering Research Center for Miniaturized Detection System, College of Life Science, Northwest University, Xi’an 710069, China
  • Received:2006-10-15 Revised:2006-11-21 Online:2007-05-30 Published:1985-09-25

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摘要：

建立了一种用Ni2+螯合的Chelating Sepharose Fast Flow亲和柱色谱和Sephadex G-75凝胶排阻柱色谱分离纯化重组肿瘤血管生长抑制因子Kringle 5的方法。采用该工艺得到的重组Kringle 5经十二烷基硫酸钠-聚丙烯酰胺凝胶电泳分析表明其纯度约为98%,且具有抑制鸡胚绒毛尿囊膜新生血管生成的生物活性。

关键词: 纯化 , 分离, 凝胶排阻柱色谱法, 亲和柱色谱法, 重组血管生长抑制因子Kringle 5

Abstract:

The Kringle 5 domain of plasminogen, which was previously shown to inhibit angiogenesis in vitro and in vivo, is one of the most potent angiogenesis inhibitors known to date. A two-step chromatographic method, which consists of Chelating Sepharose Fast Flow chelated with Ni2+ affinity medium and Sephadex G-75 medium, was established to separate and purify recombinant angiogenesis inhibitor Kringle 5 (rK5). Through the two-step chromatographic purification process, the obtained rK5 was confirmed to be homogeneous on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and its relative molecular mass was estimated to be about 32000, which matched well with the prediction by gene sequence. Its purity was about 98%, and the total protein recovery of this method was 0.63%. In addition, it was found that it inhibited the blood vessel growth of chick embryo chorioallantoic membrane effectively.

Key words: affinity column chromatography, gel exclusion column chromatography, purification , recombinant angiogenesis inhibitor Kringle 5, separation