The RecD subunit of the Escherichia coli RecBCD enzyme inhibits RecA loading, homologous recombination, and DNA repair
The RecD subunit of the Escherichia coli RecBCD enzyme inhibits RecA loading, homologous recombination, and DNA repair
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DOI：
10.1073/pnas.130192397
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发表时间：
2000-06-20
影响因子：
11.1
通讯作者：
Smith, GR
中科院分区：
文献类型：
--
作者：
Amundsen, SK;Taylor, AF;Smith, GR
RecBCD酶是大肠杆菌中同源重组和DNA修复所必需的。RecBCD酶的结构和功能在其与重组热点Chi（5 '-GCTGGTGC-3'）的相互作用上发生改变。已经假设RecD亚基在酶活性的Chi依赖性调节中起作用[Thaler，D.美国，Sampson，E.，西迪基岛罗森伯格S. M.，斯塔尔。F. W. &斯塔尔，M.（1988）在DNA损伤处理的机制和后果，编辑。Friedberg，E. & Hanawalt。P.（利斯，纽约），pp. 413-422;丘吉尔，J. J.，安德森。D. G. & Kowalczykowski，S. C.（1999）Genes Dev. 13，901-911]。我们测试的假设，RecD亚基抑制重组删除recD从核酸酶和重组缺陷型突变体recB（D1080）ACD。我们在这里报告，所产生的菌株，recB（D1080 A）C。擅长重组和DNA修复纯化能力伴随着酶活性的变化：RecB（D1080 A）C酶在DNA解旋过程中将RecA蛋白加载到DNA上，而RecB（D1080 A）CD酶则没有。总之，这些遗传和生物化学结果表明，RecBCD酶加载RecA是在E.大肠杆菌细胞，并建议RecD干扰其加载所需的酶结构域。一个核酸酶依赖的信号似乎是需要在RecD的变化，允许RecA加载。由于野生型RecBCD酶在Chi位点起作用之前未观察到RecA加载，因此我们的观察结果支持RecD抑制重组直到酶作用于Chi的观点。
The RecBCD enzyme is required for homologous recombination and DNA repair in Escherichia coli. The structure and function of RecBCD enzyme is altered on its interaction with the recombination hotspot Chi (5'-GCTGGTGC-3'). It has been hypothesized that the RecD subunit plays a role in Chi-dependent regulation of enzyme activity [Thaler, D. S., Sampson, E., Siddiqi, I., Rosenberg. S. M., stahl. F. W. & Stahl, M. (1988) in Mechanisms and Consequences of DNA Damage Processing, eds. Friedberg, E. & Hanawalt. P. (Liss, New York), pp. 413-422; Churchill, J. J., Anderson. D. G. & Kowalczykowski, S. C. (1999) Genes Dev. 13, 901-911]. We tested the hypothesis that the RecD subunit inhibits recombination by deleting recD from the nuclease- and recombination-deficient mutant recB(D1080)ACD. We report here that the resulting strain, recB(D1080A)C. was proficient for recombination and DNA repair. Recombination proficiency was accompanied by a change in enzyme activity: RecB(D1080A)C enzyme loaded RecA protein onto DNA during DNA unwinding whereas RecB(D1080A)CD enzyme did not. Together, these genetic and biochemical results demonstrate that RecA loading by RecBCD enzyme is required for recombination in E. coli cells and suggest that RecD interferes with the enzyme domain required for its loading. A nuclease-dependent signal appears to be required for a change in RecD that allows RecA loading. Because RecA loading is not observed with wild-type RecBCD enzyme until it acts at a Chi site, our observations support the view that RecD inhibits recombination until the enzyme acts at Chi.