Leite Wellington C, Galvão Carolina W, Saab Sérgio C, Iulek Jorge, Etto Rafael M, Steffens Maria B R, Chitteni-Pattu Sindhu, Stanage Tyler, Keck James L, Cox Michael M
Department of Physics, Ponta Grossa State University (UEPG), Av. Carlos Cavalcanti, 4748, CEP. 84.030-900, Ponta Grossa, PR, Brazil.
Department of Structural and Molecular Biology and Genetics, Ponta Grossa State University (UEPG), CEP 84030-900, Ponta Grossa, PR, Brazil.
PLoS One. 2016 Jul 22;11(7):e0159871. doi: 10.1371/journal.pone.0159871. eCollection 2016.
The bacterial RecA protein plays a role in the complex system of DNA damage repair. Here, we report the functional and structural characterization of the Herbaspirillum seropedicae RecA protein (HsRecA). HsRecA protein is more efficient at displacing SSB protein from ssDNA than Escherichia coli RecA protein. HsRecA also promotes DNA strand exchange more efficiently. The three dimensional structure of HsRecA-ADP/ATP complex has been solved to 1.7 Å resolution. HsRecA protein contains a small N-terminal domain, a central core ATPase domain and a large C-terminal domain, that are similar to homologous bacterial RecA proteins. Comparative structural analysis showed that the N-terminal polymerization motif of archaeal and eukaryotic RecA family proteins are also present in bacterial RecAs. Reconstruction of electrostatic potential from the hexameric structure of HsRecA-ADP/ATP revealed a high positive charge along the inner side, where ssDNA is bound inside the filament. The properties of this surface may explain the greater capacity of HsRecA protein to bind ssDNA, forming a contiguous nucleoprotein filament, displace SSB and promote DNA exchange relative to EcRecA. Our functional and structural analyses provide insight into the molecular mechanisms of polymerization of bacterial RecA as a helical nucleoprotein filament.
细菌RecA蛋白在复杂的DNA损伤修复系统中发挥作用。在此，我们报道了巴西固氮螺菌RecA蛋白（HsRecA）的功能和结构特征。HsRecA蛋白从单链DNA上取代单链结合蛋白（SSB）的效率比大肠杆菌RecA蛋白更高。HsRecA还能更有效地促进DNA链交换。HsRecA-ADP/ATP复合物的三维结构已解析到1.7 Å分辨率。HsRecA蛋白包含一个小的N端结构域、一个中央核心ATP酶结构域和一个大的C端结构域，这些结构域与同源细菌RecA蛋白相似。比较结构分析表明，古细菌和真核生物RecA家族蛋白的N端聚合基序也存在于细菌RecA蛋白中。根据HsRecA-ADP/ATP的六聚体结构重建静电势，结果显示沿着内侧有高正电荷，单链DNA在细丝内部结合于此。该表面的性质可能解释了HsRecA蛋白相对于EcRecA在结合单链DNA、形成连续核蛋白细丝、取代SSB以及促进DNA交换方面具有更大能力的原因。我们的功能和结构分析为细菌RecA作为螺旋核蛋白细丝聚合的分子机制提供了见解。
