Kim J I, Cox M M, Inman R B
Department of Biochemistry, College of Agricultural and Life Sciences, University of Wisconsin, Madison 53706.
J Biol Chem. 1992 Aug 15;267(23):16438-43.
RecA protein promotes a substantial DNA strand exchange reaction in the presence of adenosine 5'-O-3-(thio)triphosphate (ATP gamma S) (Menetski, J.P., Bear, D.G., and Kowalczykowski, S.C. (1990) Proc. Natl. Acad. Sci. U.S.A. 87, 21-25), calling into question the role of ATP hydrolysis in the strand exchange reaction. Here, we demonstrate that the ATP gamma S-mediated reaction can go to completion when the duplex DNA substrate is only 1.3 kilobase pairs in length. The ATP gamma S-mediated reaction, however, is completely blocked by a 52-base pair heterologous insertion in either DNA substrate. This same barrier is readily bypassed when ATP replaces ATP gamma S. This indicates that at least one function of recA-mediated ATP hydrolysis is to bypass structural barriers in one or both DNA substrates during strand exchange. This suggests that ATP hydrolysis is directly coupled to the branch migration phase of strand exchange, not to promote strand exchange between homologous DNA substrates during recombination, but instead to facilitate the bypass of structural barriers likely to be encountered during recombinational DNA repair.
在5'-O-3-(硫代)三磷酸腺苷(ATPγS)存在的情况下，RecA蛋白能促进大量的DNA链交换反应（梅内茨基，J.P.，贝尔，D.G.，和科瓦尔奇科夫斯基，S.C.（1990年）《美国国家科学院院刊》87卷，21 - 25页），这使得ATP水解在链交换反应中的作用受到质疑。在此，我们证明当双链DNA底物长度仅为1.3千碱基对时，ATPγS介导的反应能够完成。然而，ATPγS介导的反应在任何一个DNA底物中被一个52碱基对的异源插入完全阻断。当ATP取代ATPγS时，这个相同的障碍很容易被绕过。这表明RecA介导的ATP水解的至少一个功能是在链交换过程中绕过一个或两个DNA底物中的结构障碍。这表明ATP水解直接与链交换的分支迁移阶段相关联，不是为了促进重组过程中同源DNA底物之间的链交换，而是为了便于绕过重组性DNA修复过程中可能遇到的结构障碍。
