猪眼晶体钙结合蛋白的分离、纯化及其性质的研究

用疏水性苯基琼脂糖、Sephacryl S—200和DEAE纤维素柱层析从猪眼晶体中提纯了五种钙结合蛋白(CaBP)。DEAE纤维素柱采用了两种不同梯度的洗脱体系。纯化的CaBP的分子量子别为11,000、12,000、18,000、72,000和87,000,故分别称之为CaBP_(11)、CaBP_(12)、CaM、CaBP_(72)和CaBP_(87)。在SDS—PAGE中,除CaBP_(72)显示出三条不同区带外,其他均为单一区带。晶体CaM对环核苷酸磷酸二酯酶(PDE)的刺激作用与牛脑和猪脑CaM相同。CaBP_(11);CaBP_(12)对CaBP_(72)抗体的交叉反应率小于0.1％,但CaBP_(11)和CaBP_(12)的混合物与CaBP_(72)抗体的交叉反应率明显增加。晶体CaM可与兔抗猪脑CaM抗体1:1结合。

THE PURIFICATION AND PROPERTIES OF CALCIUM BINDING PROTEIN IN PIG LENS

Flve calcium binding proteins were separated from pig lens extracts by phenyl-Sepharose affinity chromatography, and further purified by Se-phacryl S-200 and DEAE cellulose column chromatography. Two different gradient elutions were used in DEAE cellulose column. The molecular weights of purified CaBP are 11,000, 12,000, 18,000,72,000 and 87, 000. The CaBP72(M. W. 72,000) showed three bands in SDS-PAGE, while the other CaBPs are all single bands.The lens CaM(M.W. 18,000) stimulated PDE activity in the same manner as the bovine or porcine brain calmodulin. The cross-reaction of CaBP11 or CaBP12 with anti-CaBP72 antibody was less than 0.1%, but the cross-reaction of the mixed CaBP11 and CaBP 12 with the antibody was sinifican-tly increased. Len CaM bound to rabbit anti-pig brain CaM antibody in the ratio of 1 : 1.