Mezzina M, Franchi E, Izzo R, Bertazzoni U, Rossignol J M, Sarasin A
Biochem Biophys Res Commun. 1985 Nov 15;132(3):857-63. doi: 10.1016/0006-291x(85)91886-8.
Using a method that detects catalytically active DNA ligase in NaDodSO4-polyacrylamide gels (activity gels) we have characterized ligase produced in CV1-P monkey kidney cells infected with SV40 or treated with mitomycin C. Purification on hydroxylapatite columns of DNA ligase from control cells results in two peaks of activity called ligases I and II, respectively. Analysis of ligase I on activity gels revealed major catalytic peptides with Mr of 120, 110, 70 and 58 kDa, while analysis of ligase II revealed two major peptides of 65 and 58 kDa. Infecting CV1-P cells with SV40 produced a significant increase in the 120, 110, 70 and 58 kDa peptides while treating them with mitomycin C produced a significant increase in the 70 and 58 kDa peptides and a decrease in the 120 and 110 kDa ones. Autoproteolysis of partially purified ligase under several conditions resulted in an increase in the 58 kDa peptide and in the disappearance of other peptides. These results suggest that at least one active polypeptide is common to ligases I and II.
我们采用一种能在十二烷基硫酸钠-聚丙烯酰胺凝胶（活性凝胶）中检测具有催化活性的DNA连接酶的方法，对感染了SV40或用丝裂霉素C处理的CV1-P猴肾细胞中产生的连接酶进行了特性分析。从对照细胞的DNA连接酶在羟基磷灰石柱上进行纯化，结果得到两个活性峰，分别称为连接酶I和连接酶II。在活性凝胶上对连接酶I进行分析，发现主要的催化肽的分子量分别为120、110、70和58 kDa，而对连接酶II进行分析时，发现了分子量为65和58 kDa的两个主要肽段。用SV40感染CV1-P细胞后，120、110、70和58 kDa的肽段显著增加，而用丝裂霉素C处理这些细胞后，70和58 kDa的肽段显著增加，120和110 kDa的肽段减少。在几种条件下对部分纯化的连接酶进行自催化水解，导致58 kDa肽段增加，其他肽段消失。这些结果表明，连接酶I和连接酶II至少有一个活性多肽是相同的。
