Bianco P R, Weinstock G M
Department of Biochemistry and Molecular Biology, University of Texas Medical School, Houston 77225, USA.
Biochemistry. 1998 May 19;37(20):7313-20. doi: 10.1021/bi9731293.
The RecA protein of Escherichia coli is required for homologous genetic recombination and induction of the SOS regulon. In order for RecA protein to function in these two roles, a nucleoside triphosphate cofactor, usually ATP or dATP, is required. We have examined the ability of UTP to substitute for (r,d)ATP as nucleoside triphosphate cofactor. We have found that although UTP is hydrolyzed by RecA protein in the presence of long DNA molecules, it is not hydrolyzed in reactions in which the cofactors are oligodeoxyribonucleotides less than approximately 50 nt in length. We show that UTP can efficiently substitute for ATP as nucleoside triphosphate cofactor for the DNA strand exchange reaction in vitro. The RecA1332 protein (Cys129 --> Met), which was originally shown to be defective for homologous recombination in vivo, is able to perform DNA strand exchange in vitro with ATP, but is unable to do so with UTP. These results suggest that UTP may be a cofactor for DNA strand exchange in vivo. The inability of RecA protein to hydrolyze UTP with oligodeoxyribonucleotides as cofactor and the ability of RecA to utilize UTP as cofactor in DNA strand exchange suggest a separation of the functions of RecA protein into those that require exclusively ATP and those which can utilize additional nucleoside triphosphate cofactors.
大肠杆菌的RecA蛋白对于同源基因重组和SOS调节子的诱导是必需的。为了使RecA蛋白在这两种作用中发挥功能，需要一种核苷三磷酸辅因子，通常是ATP或dATP。我们研究了UTP替代(r,d)ATP作为核苷三磷酸辅因子的能力。我们发现，尽管在长DNA分子存在的情况下UTP会被RecA蛋白水解，但在辅因子为长度小于约50个核苷酸的寡脱氧核糖核苷酸的反应中它不会被水解。我们表明，在体外DNA链交换反应中，UTP可以有效地替代ATP作为核苷三磷酸辅因子。RecA1332蛋白（Cys129→Met）最初被证明在体内同源重组方面存在缺陷，它能够在体外与ATP进行DNA链交换，但不能与UTP进行。这些结果表明UTP可能是体内DNA链交换的一种辅因子。RecA蛋白不能以寡脱氧核糖核苷酸作为辅因子水解UTP，以及RecA在DNA链交换中能够利用UTP作为辅因子，这表明RecA蛋白的功能分为那些仅需要ATP的功能和那些可以利用额外核苷三磷酸辅因子的功能。
