Base excision repair (BER) ensures correction of most abundant DNA lesions in mammals. The efficiency of this multistep DNA repair process that can occur via different pathways depends on the coordinated action of enzymes catalyzing its individual steps. The scaffold XRCC1 (X-ray repair cross-complementing protein 1) protein plays an important coordinating role in the repair of damaged bases and apurinic/apyrimidinic (AP) sites via short-patch (SP) BER pathway, as well as in the repair of single-strand DNA breaks. In this study, we demonstrated for the first time in vitro formation of the ternary XRCC1 complex with the key enzymes of SP BER — DNA polymerase β (Polβ) and DNA ligase IIIa (LiglIIa) — using the fluorescence-based technique. It was found that Polβ directly interacts with LiglIIa, but their complex is less stable than the XRCC1—Polβ and XRCC1—LigIIIa complexes. The effect of XRCC1 oxidation and composition of the multiprotein complex on the efficiency of DNA synthesis and DNA ligation during DNA repair has been explored. We found that formation of the disulfide bond between Cys12 and Cys20 residues as a result of XRCC1 oxidation (previously shown to modulate the protein affinity for Polβ), affects the yield of the final product of SP BER and of non-ligated DNA intermediates (substrates of long-patch BER). The effect of XRCC1 oxidation on the final product yield depended on the presence of AP endonuclease 1. Together with the data from our previous work, the results of this study suggest an important role of XRCC1 oxidation in the fine regulation of formation of BER complexes and their functional activity.

中文翻译：

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人 XRCC1 蛋白氧化对其与 DNA 碱基切除修复关键酶复合物功能活性的影响

碱基切除修复 (BER) 可确保纠正哺乳动物中最丰富的 DNA 损伤。这种可以通过不同途径发生的多步 DNA 修复过程的效率取决于催化其各个步骤的酶的协调作用。支架 XRCC1（X 射线修复交叉互补蛋白 1）蛋白在通过短补丁 (SP) BER 途径修复受损碱基和无嘌呤/无嘧啶 (AP) 位点以及修复中发挥重要的协调作用单链 DNA 断裂。在这项研究中，我们首次使用基于荧光的技术证明了三元 XRCC1 复合物与 SP BER 的关键酶 DNA 聚合酶 β (Polβ) 和 DNA 连接酶 IIIa (LiglIIa) 的体外形成。发现 Polβ 直接与 LiglIIa 相互作用，但它们的复合物不如 XRCC1-Polβ 和 XRCC1-LigIIIa 复合物稳定。已经探索了 XRCC1 氧化和多蛋白复合物的组成对 DNA 修复过程中 DNA 合成和 DNA 连接效率的影响。我们发现，由于 XRCC1 氧化（之前显示可调节蛋白质对 Polβ 的亲和力），Cys12 和 Cys20 残基之间二硫键的形成会影响 SP BER 和非连接 DNA 中间体（底物）的最终产物的产量长补丁 BER）。XRCC1 氧化对最终产物产率的影响取决于 AP 核酸内切酶 1 的存在。与我们之前工作的数据一起，本研究的结果表明 XRCC1 氧化在精细调节 BER 复合物和他们的功能活动。