The scaffold protein XRCC1 stabilizes the formation of polβ/gap DNA and ligase IIIα/nick DNA complexes in base excision repair.
The scaffold protein XRCC1 stabilizes the formation of polβ/gap DNA and ligase IIIα/nick DNA complexes in base excision repair.
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DOI：
10.1016/j.jbc.2021.101025
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发表时间：
2021-09
期刊：
影响因子：
--
通讯作者：
Çağlayan M
中科院分区：
文献类型：
--
作者：
Tang Q;Çağlayan M
碱基切除修复（BER）途径涉及DNA聚合酶（pol）β的缺口填充和随后连接酶IIIα的缺口封闭。X射线交叉互补蛋白1（XRCC 1），一种非酶支架蛋白，组装多蛋白复合物，尽管XRCC 1协调协调BER的最后步骤的机制仍然不完全确定。在这里，使用生物化学和生物物理方法的组合，我们揭示了polβ/XRCC 1复合物在正确的核苷酸插入DNA中的缺口后增加BER反应的持续合成能力，并增强了切口修复产物向最终连接步骤的传递。此外，在XRCC 1存在下，polβ 8-oxodGTP插入后切口修复中间体的诱变连接增强。结果表明，XRCC 1对polβ/dNTP/gap DNA和连接酶Ⅲ α/ATP/nick DNA催化三元复合物的形成具有稳定作用。蛋白质-蛋白质相互作用和DNA结合动力学的实时监测显示，XRCC 1与polβ的结合比与连接酶IIIα或aprataxin的结合更强，并且与未受损或受损末端的切口DNA的亲和力比与一个核苷酸缺口修复中间体的亲和力更高。最后，我们证明了稳定的polβ/XRCC 1复合物形成、polβ和连接酶IIIα蛋白相互作用动力学以及作为癌症相关（P161 L、R194 W、R280 H、R399 Q、Y 576 S）和小脑共济失调相关（K431 N）XRCC 1变体的结果的交接过程的轻微差异。总的来说，我们的研究结果为XRCC 1的协调作用及其疾病相关变体对多蛋白/DNA复合物中底物-产物通道的影响提供了新的见解，以实现有效的BER。
The base excision repair (BER) pathway involves gap filling by DNA polymerase (pol) β and subsequent nick sealing by ligase IIIα. X-ray cross-complementing protein 1 (XRCC1), a nonenzymatic scaffold protein, assembles multiprotein complexes, although the mechanism by which XRCC1 orchestrates the final steps of coordinated BER remains incompletely defined. Here, using a combination of biochemical and biophysical approaches, we revealed that the polβ/XRCC1 complex increases the processivity of BER reactions after correct nucleotide insertion into gaps in DNA and enhances the handoff of nicked repair products to the final ligation step. Moreover, the mutagenic ligation of nicked repair intermediate following polβ 8-oxodGTP insertion is enhanced in the presence of XRCC1. Our results demonstrated a stabilizing effect of XRCC1 on the formation of polβ/dNTP/gap DNA and ligase IIIα/ATP/nick DNA catalytic ternary complexes. Real-time monitoring of protein–protein interactions and DNA-binding kinetics showed stronger binding of XRCC1 to polβ than to ligase IIIα or aprataxin, and higher affinity for nick DNA with undamaged or damaged ends than for one nucleotide gap repair intermediate. Finally, we demonstrated slight differences in stable polβ/XRCC1 complex formation, polβ and ligase IIIα protein interaction kinetics, and handoff process as a result of cancer-associated (P161L, R194W, R280H, R399Q, Y576S) and cerebellar ataxia-related (K431N) XRCC1 variants. Overall, our findings provide novel insights into the coordinating role of XRCC1 and the effect of its disease-associated variants on substrate-product channeling in multiprotein/DNA complexes for efficient BER.
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影响因子：
14.9
作者：
Berquist BR;Singh DK;Fan J;Kim D;Gillenwater E;Kulkarni A;Bohr VA;Ackerman EJ;Tomkinson AE;Wilson DM 3rd
通讯作者：
Wilson DM 3rd
影响因子：
3.8
作者：
Çağlayan M
通讯作者：
Çağlayan M
影响因子：
7.4
作者：
Abbotts R;Wilson DM 3rd
通讯作者：
Wilson DM 3rd
影响因子：
3.8
作者：
Sizova, Dania V.;Keh, Agnes;Taylor, Ben F.;Sweasy, Joann B.
通讯作者：
Sweasy, Joann B.
影响因子：
4.8
作者：
Beernink, PT;Hwang, M;Thelen, MP
通讯作者：
Thelen, MP