DNA Polymerase Gp90 Activities and Regulations on Strand Displacement DNA Synthesis Revealed at Single-molecule Level
DNA Polymerase Gp90 Activities and Regulations on Strand Displacement DNA Synthesis Revealed at Single-molecule Level
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DNA 聚合酶 Gp90 活性和对链置换 DNA 合成的调节在单分子水平上揭示
DOI：
10.1096/fj.202100033rr
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发表时间：
2021
期刊：
影响因子：
--
通讯作者：
Huidong Zhang
中科院分区：
文献类型：
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作者：
Shuming Zhang;Xue Xiao;Jingwei Kong;Ke Lu;Shuo-Xing Dou;Peng-Ye Wang;Lu Ma;Yuru Liu;Guohong Li;Wei Li;Huidong Zhang
链置换DNA合成(SDDS)是DNA复制中必不可少的一步。我们用磁钳在单分子水平上研究了野生型gp90及其外切酶缺失的gp90外切酶的SDDS动力学。在SDDS之前确认了gp90与叉状瓣的一种新的结合状态，这表明在SDDS的启动过程中存在一个中间体。Gp90外源或wt-gp90对SDDS的反应速率和加工率随力和dNTP浓度的增加而增加。Wt-gp90的核酸外切速度和加工率随力的降低而降低。高GC含量会降低wt-gp90的SDDS和核酸外切酶的加工率，但会增加外切酶的加工率。较高的外切力和dNTP浓度以及较低的GC含量促进了连续的SDDS，但延缓了wt-gp90的连续核酸外切酶。此外，增加GC含量会加速SDDS或核酸外切酶向核酸外切酶的转变。这项工作详细揭示了SDDS的动力学，并在单聚合酶水平上对各种因素对SDDS的调节提供了更广泛的认识。
Strand displacement DNA synthesis (SDDS) is an essential step in DNA replication. With magnetic tweezers, we investigated SDDS kinetics of wild‐type gp90 and its exonuclease‐deficient polymerase gp90 exo‐at single‐molecule level. A novel binding state of gp90 to the fork flap was confirmed prior to SDDS, suggesting an intermediate in the initiation of SDDS. The rate and processivity of SDDS by gp90 exo‐or wt‐gp90 are increased with force and dNTP concentration. The rate and processivity of exonuclease by wt‐gp90 are decreased with force. High GC content decreases SDDS and exonuclease processivity but increases exonuclease rate for wt‐gp90. The high force and dNTP concentration and low GC content facilitate the successive SDDS but retard the successive exonuclease for wt‐gp90. Furthermore, increasing GC content accelerates the transition from SDDS or exonuclease to exonuclease. This work reveals the kinetics of SDDS in detail and offers a broader cognition on the regulation of various factors on SDDS at single‐polymerase level.