DNA strand exchange promoted by recA protein and single-stranded DNA-binding protein of Escherichia coli.
DNA strand exchange promoted by recA protein and single-stranded DNA-binding protein of Escherichia coli.
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大肠杆菌的recA蛋白和单链DNA结合蛋白促进DNA链交换。
DOI：
10.1101/sqb.1983.047.01.092
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发表时间：
1983
期刊：
影响因子：
--
通讯作者：
Lehman,IR
中科院分区：
文献类型：
--
作者：
Cox,MM;Soltis,DA;Livneh,Z;Lehman,IR
recA 基因在大肠杆菌的同源基因重组中发挥着核心作用（Radding 1978）。它也是介导细胞对 DNA 损伤反应的调节途径中的关键元素 (Gottesman 1981)。已分离出recA基因的产物（Okawa等人，1979年；Roberts等人，1979年；Shibata等人，1979年；Weinstock等人，1979年），并且其一级结构已确定（Horii等人，1980年；Sancar等人，1980年）。与其体内两种作用相关的活性已被鉴定（Roberts et al. 1979；Radding 1981；McEntee and Weinstock 1981），并且已经开发出多种可以测量它们的测定方法（Roberts et al. 1979；Shibata et al. 1979；Weinstock et al. 1979；Cox and Lehman 1981a；West et al. 2017）。 1981a).我们的目标是确定 recA 蛋白在同源重组中的作用。我们最近的努力集中在recA蛋白促进的反应之一上，即环状（加）单链bX174 DNA（SS DNA）和同源线性双链体bX174 DNA之间的链交换与ATP水解为ADP和P相结合（Cox和Lehman 1981a，b，1982；Cox等人1982a，b）（图1）。 1）。该反应在体外模拟同源重组的步骤，其中recA蛋白很可能在体内参与。它具有几个重要的优点：(1) 底物和产物均具有良好的特征且易于区分；(2) 可以使用多种测定法，包括允许直接测量异源双链体形成的测定法； (3)反应高效，简化了动力学分析。链交换的效率取决于大肠杆菌单链 DNA 结合蛋白 (SSB) 的存在。事实上，有遗传证据表明 SSB 可能在重组和重组修复中发挥作用（Johnson 1977；Glassberg 等人 1979；Whittier 和 Chase 1981）。除了上述三链交换之外，recA 蛋白还将促进涉及四个 DNA 链的相互交换（West 等人，1981a；Das-Gupta 等人，1981）。 West等人（1982）最近表明，SSB虽然刺激三链交换，但对四链交换没有影响或轻微抑制。需要更多的工作来了解这两种类型的交换与体内同源重组的关系。我们最初的努力是通过
The recA gene plays a central role in homologous genetic recombination in Escherichia coli (Radding 1978). It is also a key element in a regulatory pathway mediating cellular responses to DNA damage (Gottesman 1981). The product of the recA gene has been isolated (Ogawa et al. 1979; Roberts et al. 1979; Shibata et al. 1979; Weinstock et al. 1979), and its primary structure has been determined (Horii et al. 1980; Sancar et al. 1980). Activities relevant to both of its roles in vivo have been identified (Roberts et al. 1979; Radding 1981; McEntee and Weinstock 1981), and a variety of assays have been developed with which they can be measured (Roberts et al. 1979; Shibata et al. 1979; Weinstock et al. 1979; Cox and Lehman 1981a; West et al. 1981a).Our aim is to determine the mechanism of action of recA protein in homologous recombination. Our recent efforts have concentrated on one of the recA-protein-promoted reactions, the exchange of strands between circular (plus) single-stranded~ bX174 DNA (SS DNA) and homologous linear duplex~ bX174 DNA coupled to the hydrolysis of ATP to ADP and P,(Cox and Lehman 1981a, b, 1982; Cox et al. 1982a, b)(Fig. 1). This reaction mimics, in vitro, steps in homologous recombination in which recA protein very likely participates in vivo. It has several important advantages:(1) both substrates and products are well characterized and easily distinguishable;(2) a variety of assays are available, including one that permits the direct measurement of heteroduplex formation; and (3) the reaction is efficient, thereby simplifying kinetic analysis. The efficiency of strand exchange is dependent on the presence of the single-stranded DNA-binding protein (SSB) of E. coli. There is, in fact, genetic evidence that SSB may play a role in recombination and recombinational repair (Johnson 1977; Glassberg et al. 1979; Whittier and Chase 1981). In addition to the three-strand exchange described above, recA protein will promote a reciprocal exchange involving four DNA strands (West et al. 1981a; Das-Gupta et al. 1981). West et al.(1982) have recently shown that SSB, although stimulating three-strand exchanges, either has no effect on or slightly inhibits a four-strand exchange. More work is required to understand the relationship of the two types of exchange to homologous recombination in vivo. Our initial efforts to determine the mechanism by