Both RadA and RadB Are Involved in Homologous Recombination inPyrococcus furiosus *
Both RadA and RadB Are Involved in Homologous Recombination inPyrococcus furiosus *
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DOI：
10.1074/jbc.m004557200
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发表时间：
2000-10
期刊：
影响因子：
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通讯作者：
K. Komori;T. Miyata;J. DiRuggiero;R. Holley-Shanks;I. Hayashi;I. Cann;Kota Mayanagi;H. Shinagawa-H.-Shinaga
中科院分区：
文献类型：
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作者：
K. Komori;T. Miyata;J. DiRuggiero;R. Holley-Shanks;I. Hayashi;I. Cann;Kota Mayanagi;H. Shinagawa-H.-Shinaga
RecA和Rad 51蛋白分别是细菌和真核生物同源重组所必需的。被称为RadA的同源蛋白质已被描述用于Escherichia。在这里，我们提出了两个RecA/Rad 51家族蛋白，RadA和RadB，从激烈的热球菌的特性。radA和raddB基因不受γ射线和紫外线照射以及热休克引起的DNA损伤的诱导，表明它们可能在这种超嗜热菌中组成型表达。RadA具有依赖DNA的ATP酶、D环形成和链交换活性。相比之下，RadB具有非常弱的ATP酶活性，不受DNA刺激。该蛋白对DNA有很强的结合亲和力，但几乎没有链交换活性。RadA和RadB之间的直接相互作用，通过免疫沉淀法检测。此外，RadB，而不是RadA，共沉淀与Hjc，霍利迪连接resolvase发现在P。在缺乏ATP的情况下，这种相互作用在ATP的存在下被抑制。Hjc的Holliday连接切割活性在不存在ATP的情况下被RadB抑制，但在存在ATP的情况下不被RadB抑制。这些结果表明，RadB在同源重组中起着重要作用，并可能调节分支结构DNA的切割反应。
RecA and Rad51 proteins are essential for homologous recombination in Bacteria andEukarya, respectively. Homologous proteins, called RadA, have been described for Archaea. Here we present the characterization of two RecA/Rad51 family proteins, RadA and RadB, fromPyrococcus furiosus. The radA andradB genes were not induced by DNA damage resulting from exposure of the cells to γ and UV irradiation and heat shock, suggesting that they might be constitutively expressed in this hyperthermophile. RadA had DNA-dependent ATPase, D-loop formation, and strand exchange activities. In contrast, RadB had a very weak ATPase activity that is not stimulated by DNA. This protein had a strong binding affinity for DNA, but little strand exchange activity could be detected. A direct interaction between RadA and RadB was detected by an immunoprecipitation assay. Moreover, RadB, but not RadA, coprecipitated with Hjc, a Holliday junction resolvase found inP. furiosus, in the absence of ATP. This interaction was suppressed in the presence of ATP. The Holliday junction cleavage activity of Hjc was inhibited by RadB in the absence, but not in the presence, of ATP. These results suggest that RadB has important roles in homologous recombination in Archaea and may regulate the cleavage reactions of the branch-structured DNA.