Klenow H, Henningsen I
Proc Natl Acad Sci U S A. 1970 Jan;65(1):168-75. doi: 10.1073/pnas.65.1.168.
Purification of DNA polymerase from E. coli B has in two cases each time led to the isolation of two separate polymerase activities, enzyme A and enzyme B. Enzyme A was in contrast to enzyme B almost completely devoid of exonuclease activity. Each of the two enzymes yielded a single symmetrical activity peak in gel filtration chromatograms. From the elution volumes the molecular weights were estimated to be about 70,000 for enzyme A and about 150,000 for enzyme B. Treatment of enzyme B with subtilisin led to an increase of about 30 per cent of the polymerase activity while the exonuclease activity almost completely disappeared. The product of the subtilisin treatment (enzyme C) gave rise to a single symmetrical polymerase activity peak in a gel filtration chromatogram. The elution volume was identical to that obtained with enzyme A. It is concluded that enzyme A and enzyme C are formed by limited proteolysis of enzyme B.
从大肠杆菌B中纯化DNA聚合酶，每次在两种情况下都导致分离出两种独立的聚合酶活性，即酶A和酶B。与酶B相比，酶A几乎完全没有核酸外切酶活性。两种酶在凝胶过滤色谱图中均产生一个单一的对称活性峰。根据洗脱体积估计，酶A的分子量约为70,000，酶B的分子量约为150,000。用枯草杆菌蛋白酶处理酶B导致聚合酶活性增加约30%，而核酸外切酶活性几乎完全消失。枯草杆菌蛋白酶处理的产物（酶C）在凝胶过滤色谱图中产生一个单一的对称聚合酶活性峰。洗脱体积与酶A的相同。得出的结论是，酶A和酶C是由酶B的有限蛋白水解形成的。
