Russian Academy of Sciences, Gatchina/St. Petersburg, Russia.
Mol Microbiol. 2010 Dec;78(6):1523-38. doi: 10.1111/j.1365-2958.2010.07424.x. Epub 2010 Oct 19.
The wild-type Escherichia coli RecA protein is a recombinase platform with unrealized recombination potential. We have explored the factors affecting recombination during conjugation with a quantitative assay. Regulatory proteins that affect RecA function have the capacity to increase or decrease recombination frequencies by factors up to sixfold. Autoinhibition by the RecA C-terminus can affect recombination frequency by factors up to fourfold. The greatest changes in recombination frequency measured here are brought about by point mutations in the recA gene. RecA variants can increase recombination frequencies by more than 50-fold. The RecA protein thus possesses an inherently broad functional range. The RecA protein of E. coli (EcRecA) is not optimized for recombination function. Instead, much of the recombination potential of EcRecA is structurally suppressed, probably reflecting cellular requirements. One point mutation in EcRecA with a particularly dramatic effect on recombination frequency, D112R, exhibits an enhanced capacity to load onto SSB-coated ssDNA, overcome the effects of regulatory proteins such as PsiB and RecX, and to pair homologous DNAs. Comparisons of key RecA protein mutants reveal two components to RecA recombination function - filament formation and the inherent DNA pairing activity of the formed filaments.
野生型大肠杆菌 RecA 蛋白是一种重组酶平台，具有未实现的重组潜力。我们已经通过定量测定探索了影响接合过程中重组的因素。影响 RecA 功能的调节蛋白可以将重组频率提高或降低六倍。RecA C 末端的自动抑制作用可以将重组频率提高四倍。这里测量的重组频率最大变化是由 recA 基因突变引起的。RecA 变体可以将重组频率提高 50 多倍。因此，RecA 蛋白具有固有的广泛功能范围。大肠杆菌的 RecA 蛋白（EcRecA）并非针对重组功能进行了优化。相反，EcRecA 的大部分重组潜力在结构上受到抑制，这可能反映了细胞的需求。EcRecA 中有一种突变，即 D112R，对重组频率的影响特别大，它表现出增强的能力，可加载到 SSB 包裹的 ssDNA 上，克服 PsiB 和 RecX 等调节蛋白的影响，并与同源 DNA 配对。对关键 RecA 蛋白突变体的比较揭示了 RecA 重组功能的两个组成部分 - 丝形成和形成丝的固有 DNA 配对活性。
