Zou Zhenyu, Chen Ze, Xue Qizhen, Xu Ying, Xiong Jingyuan, Yang Ping, Le Shuai, Zhang Huidong
Public Health Laboratory Sciences and Toxicology, West China School of Public Health, Sichuan University, No.17 People's South Road, Chengdu, 6100041, P. R. China.
Institute for Chemical Carcinogenesis, Guangzhou Medical University, Xinzao, Panyu District, Guangzhou, 511439, P. R. China.
Chembiochem. 2018 Jun 14. doi: 10.1002/cbic.201800203.
The DNA replisome inevitably encounters DNA damage during DNA replication. The T7 DNA replisome contains a DNA polymerase (gp5), the processivity factor thioredoxin (trx), a helicase-primase (gp4), and a ssDNA-binding protein (gp2.5). T7 protein interactions mediate this DNA replication. However, whether the protein interactions could promote DNA damage bypass is still little addressed. In this study, we investigated strand-displacement DNA synthesis past 8-oxoG or O -MeG lesions at the synthetic DNA fork by the T7 DNA replisome. DNA damage does not obviously affect the binding affinities between helicase, polymerase, and DNA fork. Relative to unmodified G, both 8-oxoG and O -MeG-as well as GC-rich template sequence clusters-inhibit strand-displacement DNA synthesis and produce partial extension products. Relative to the gp4 ΔC-tail, gp4 promotes DNA damage bypass. The presence of gp2.5 also promotes it. Thus, the interactions of polymerase with helicase and ssDNA-binding protein facilitate DNA damage bypass. Accessory proteins in other complicated DNA replisomes also facilitate bypassing DNA damage in similar manner. This work provides new mechanistic information relating to DNA damage bypass by the DNA replisome.
DNA复制体在DNA复制过程中不可避免地会遇到DNA损伤。T7 DNA复制体包含一种DNA聚合酶（gp5）、持续性因子硫氧还蛋白（trx）、一种解旋酶-引物酶（gp4）和一种单链DNA结合蛋白（gp2.5）。T7蛋白相互作用介导这种DNA复制。然而，蛋白质相互作用是否能促进DNA损伤的跨越仍鲜有人研究。在本研究中，我们研究了T7 DNA复制体在合成DNA叉处对8-氧代鸟嘌呤（8-oxoG）或O-甲基鸟嘌呤（O-MeG）损伤进行的链置换DNA合成。DNA损伤并未明显影响解旋酶、聚合酶和DNA叉之间的结合亲和力。相对于未修饰的鸟嘌呤（G），8-oxoG和O-MeG以及富含GC的模板序列簇均会抑制链置换DNA合成并产生部分延伸产物。相对于gp4的C末端缺失突变体（gp4 ΔC-tail），gp4能促进DNA损伤的跨越。gp2.5的存在也能促进这一过程。因此，聚合酶与解旋酶和单链DNA结合蛋白的相互作用有助于DNA损伤的跨越。其他复杂DNA复制体中的辅助蛋白也以类似方式促进DNA损伤的跨越。这项工作提供了有关DNA复制体跨越DNA损伤的新机制信息。
