The β2 clamp in the Mycobacterium tuberculosis DNA polymerase III αβ2ε replicase promotes polymerization and reduces exonuclease activity.
The β2 clamp in the Mycobacterium tuberculosis DNA polymerase III αβ2ε replicase promotes polymerization and reduces exonuclease activity.
复制标题
结核分枝杆菌 DNA 聚合酶 III Alpha2ε 复制酶中的 β2 钳可促进聚合并降低核酸外切酶活性。
DOI：
10.1038/srep18418
复制
发表时间：
2016-01-29
影响因子：
4.6
通讯作者：
Bi L
中科院分区：
文献类型：
--
作者：
Gu S;Li W;Zhang H;Fleming J;Yang W;Wang S;Wei W;Zhou J;Zhu G;Deng J;Hou J;Zhou Y;Lin S;Zhang XE;Bi L
DNA聚合酶III（DNA pol III）是一种多亚基复制机器，负责细菌基因组的准确和快速复制，然而，它在结核分枝杆菌（Mtb）中的功能尚需进一步研究。我们已经重建了Mtb DNA pol III全酶的体外前导链复制过程，并研究了其关键组分之间的物理和功能关系。我们通过生物化学方法和体外和体内的蛋白质-蛋白质相互作用测定验证了αβ2ε聚合酶-钳-核酸外切酶复制酶复合物的存在，并证实除了其α亚基的聚合酶活性外，Mtb DNA pol III具有两个潜在的校正亚基：α和ε亚基。在DNA复制过程中，β2夹的存在强烈促进αβ2ε复制酶的聚合并降低其核酸外切酶活性。我们的工作为进一步研究复制机制在复制和校对之间切换的机制提供了基础，并为选择靶向Mtb DNA复制的抗菌药物提供了实验平台。
DNA polymerase III (DNA pol III) is a multi-subunit replication machine responsible for the accurate and rapid replication of bacterial genomes, however, how it functions in Mycobacterium tuberculosis (Mtb) requires further investigation. We have reconstituted the leading-strand replication process of the Mtb DNA pol III holoenzyme in vitro, and investigated the physical and functional relationships between its key components. We verify the presence of an αβ2ε polymerase-clamp-exonuclease replicase complex by biochemical methods and protein-protein interaction assays in vitro and in vivo and confirm that, in addition to the polymerase activity of its α subunit, Mtb DNA pol III has two potential proofreading subunits; the α and ε subunits. During DNA replication, the presence of the β2 clamp strongly promotes the polymerization of the αβ2ε replicase and reduces its exonuclease activity. Our work provides a foundation for further research on the mechanism by which the replication machinery switches between replication and proofreading and provides an experimental platform for the selection of antimicrobials targeting DNA replication in Mtb.