Department of Biochemistry and Biomedical Sciences and M. G. DeGroote Institute for Infectious Disease Research, McMaster University, 1200 Main Street West, Hamilton, Ontario L8N 3Z5, Canada and Laboratory of Molecular Biology, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, 5 Center Drive, Bethesda, MD 20892, USA.
Nucleic Acids Res. 2013 Nov;41(21):9934-44. doi: 10.1093/nar/gkt759. Epub 2013 Aug 23.
The ability of Deinococcus radiodurans to recover from extensive DNA damage is due in part to its ability to efficiently repair its genome, even following severe fragmentation by hundreds of double-strand breaks. The single-strand annealing pathway plays an important role early during the recovery process, making use of a protein, DdrB, shown to greatly stimulate ssDNA annealing. Here, we report the structure of DdrB bound to ssDNA to 2.3 Å. Pentameric DdrB was found to assemble into higher-order structures that coat ssDNA. To gain further mechanistic insight into the protein's function, a number of point mutants were generated altering both DNA binding and higher order oligomerization. This work not only identifies higher-order DdrB associations but also suggests the presence of an extended DNA binding surface running along the 'top' surface of a DdrB pentamer and continuing down between two individual subunits of the ring structure. Together this work sheds new insight into possible mechanisms for DdrB function in which higher-order assemblies of DdrB pentamers assist in the pairing of complementary ssDNA using an extended DNA binding surface.
耐辐射球菌能够从广泛的 DNA 损伤中恢复，部分原因是它能够有效地修复基因组，即使在数百个双链断裂严重碎片化后也是如此。单链退火途径在恢复过程的早期起着重要作用，利用一种称为 DdrB 的蛋白质来极大地促进 ssDNA 退火。在这里，我们报告了 DdrB 与 ssDNA 结合的结构，分辨率为 2.3Å。五聚体 DdrB 被发现组装成覆盖 ssDNA 的更高阶结构。为了更深入地了解蛋白质的功能，我们生成了许多点突变体，改变了 DNA 结合和高阶寡聚化。这项工作不仅确定了 DdrB 的高阶关联，还表明存在一个沿 DdrB 五聚体“顶部”表面延伸的 DNA 结合表面，并在环结构的两个单独亚基之间延伸。这项工作为 DdrB 功能的可能机制提供了新的见解，其中 DdrB 五聚体的高阶组装有助于使用扩展的 DNA 结合表面对互补 ssDNA 进行配对。
