Institute of Medical Science and Technology, Kaohsiung, Taiwan; Department of Chemistry, Kaohsiung, Taiwan; Aerosol Science Research Center, National Sun Yat-sen University, Kaohsiung, Taiwan.
Department of Life Sciences and Institute of Genome Sciences, National Yang-Ming University, Taipei, Taiwan.
Biophys J. 2021 Aug 3;120(15):3166-3179. doi: 10.1016/j.bpj.2021.06.004. Epub 2021 Jun 29.
The C-terminus of Escherichia coli RecA protein can affect the DNA binding affinity, interact with accessory proteins, and regulate the RecA activity. A substantial upward shift in the pH-reaction profile of RecA-mediated DNA strand-exchange reactions was observed for C-terminal-truncated E. coli ΔC17 RecA, Deinococcus radiodurans RecA, and Deinococcus ficus RecA. Here, the process of RecA-mediated strand exchange from the beginning to the end was investigated with florescence resonance energy transfer and tethered particle motion experiments to determine the detailed regulation mechanism. RecA proteins with a shorter C-terminus possess more stable nuclei, higher DNA binding affinities, and lower protonation requirements for the formation of nucleoprotein filaments. Moreover, more stable synaptic complexes in the homologous sequence searching process were also observed for RecA proteins with a shorter C-terminus. Our results suggest that the C-terminus of RecA proteins regulates not only the formation of RecA nucleoprotein filaments but also the entrance of secondary DNA into RecA nucleoprotein filaments.
大肠杆菌 RecA 蛋白的 C 端可以影响 DNA 结合亲和力，与辅助蛋白相互作用，并调节 RecA 活性。对于 C 端截断的大肠杆菌 ΔC17 RecA、耐辐射球菌 RecA 和榕生球菌 RecA，观察到 RecA 介导的 DNA 链交换反应的 pH 反应谱有显著向上移动。在这里，通过荧光共振能量转移和束缚粒子运动实验从开始到结束研究 RecA 介导的链交换过程，以确定详细的调节机制。C 端较短的 RecA 蛋白具有更稳定的核、更高的 DNA 结合亲和力和形成核蛋白丝所需的更低质子化要求。此外，对于 C 端较短的 RecA 蛋白，在同源序列搜索过程中也观察到更稳定的突触复合物。我们的结果表明，RecA 蛋白的 C 端不仅调节 RecA 核蛋白丝的形成，而且调节二级 DNA 进入 RecA 核蛋白丝。
