Yoshida H, Murachi T, Tsukahara I
Biochim Biophys Acta. 1984 Apr 10;798(2):252-9. doi: 10.1016/0304-4165(84)90313-1.
A Ca2+-dependent cysteine proteinase (calpain, EC 3.4.22.17) was found in the cystosolic fraction of bovine lens and purified to apparent homogeneity. The purified enzyme required 1 mM Ca2+ for its full activation and was composed of two subunits of Mr 80 000 and 29 000 as demonstrated by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate (SDS). This enzyme, when activated by Ca2+, degraded both A- and B-chains of alpha-crystallin, which were isolated also from bovine lens. SDS-gel electrophoresis of the digest revealed that the A-chain (Mr 19 500) was broken down to produce an 18-kDa polypeptide fragment and the B-chain (Mr 22 500) to produce a 19.5-kDa polypeptide fragment. No further cleavage occurred even upon prolonged incubation or after the second addition of the enzyme, indicating the uniquely limited proteolysis of each chain protein. The existence of calpastatin, an endogenous inhibitor protein specific for calpain, was also demonstrated in bovine lens cytosol.
在牛晶状体的胞质组分中发现了一种钙依赖性半胱氨酸蛋白酶（钙蛋白酶，EC 3.4.22.17），并将其纯化至表观均一性。纯化后的酶完全活化需要1 mM Ca2+，在十二烷基硫酸钠（SDS）存在下进行聚丙烯酰胺凝胶电泳显示，该酶由分子量为80 000和29 000的两个亚基组成。这种酶在被Ca2+激活后，会降解同样从牛晶状体中分离出来的α-晶状体蛋白的A链和B链。消化产物的SDS-凝胶电泳显示，A链（Mr 19 500）被分解产生一个18 kDa的多肽片段，B链（Mr 22 500）被分解产生一个19.5 kDa的多肽片段。即使长时间孵育或再次添加该酶后，也不会发生进一步的切割，这表明每条链蛋白的蛋白水解作用具有独特的局限性。在牛晶状体胞质溶胶中也证实了钙蛋白酶抑制蛋白（一种对钙蛋白酶具有特异性的内源性抑制蛋白）的存在。
