Stable binding of human XPC complex to irradiated DNA confers strong discrimination for damaged sites
Stable binding of human XPC complex to irradiated DNA confers strong discrimination for damaged sites
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DOI：
10.1006/jmbi.2000.3857
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发表时间：
2000-07-07
影响因子：
5.6
通讯作者：
Wood, RD
中科院分区：
文献类型：
--
作者：
Batty, D;Rapic'-Otrin, V;Wood, RD
DNA损伤的核苷酸切除修复（NER）需要一种有效的手段来区分受损和未受损的DNA。来自患有着色性干皮病C组的人的细胞在基因组的大部分中不执行NER，并且通过与hHR 23 B蛋白形成复合物的XPC蛋白进行校正。这种复合物优先与某些类型的受损DNA结合，但与其他NER蛋白质相比的区分程度尚不清楚。纯化重组XPC、hHR 23 B和XPC-hHR 23 B复合物。在重建的修复系统中，hHR 23 B刺激XPC活性十倍。电泳迁移率变化竞争测量揭示了XPC-hHR 23 B与UV损伤的DNA结合的优先性是未损伤DNA的400倍。这种损伤偏好比XPA蛋白所显示的要大得多。尽管UV-DDB对DNA的摩尔亲和力大得多，但鉴别能力与本文平行测定的XP-E因子UV-DDB相似。XPC-hHR 23 B与UV损伤的DNA的结合非常快。损伤的DNA-XPC-hHR 23 B复合物是稳定的，在30 ℃下用过量的W-损伤的DNA攻击后，一半的复合物保留了4小时。XPC-hHR 23 B对（6-4）光产物的亲和力水平高于环丁烷嘧啶二聚体，并且对用顺铂和烷化剂处理的DNA具有一定的亲和力。XPC-hHR 23 B可以与单链M13 DNA结合，但与单链均聚物的结合很差。XPC复合物对受损双链DNA结构的强烈偏好表明其在人类NER期间作为非转录DNA中的主要损伤识别因子的重要性。(C)北京大学出版社.
Nucleotide excision repair (NER) of DNA damage requires an efficient means of discrimination between damaged and non-damaged DNA. Cells from humans with xeroderma pigmentosum group C do not perform NER in the bulk of the genome and are corrected by XPC protein, which forms a complex with hHR23B protein. This complex preferentially binds to some types of damaged DNA, but the extent of discrimination in comparison to other NER proteins has not been clear. Recombinant XPC, hHR23B, and XPC-hHR23B complex were purified. In a reconstituted repair system, hHR23B stimulated XPC activity tenfold. Electrophoretic mobility-shift competition measurements revealed a 400-fold preference for binding of XPC-hHR23B to UV damaged over non-damaged DNA. This damage preference is much greater than displayed by the XPA protein. The discrimination power is similar to that determined here in parallel for the XP-E factor UV-DDB, despite the considerably greater molar affinity of UV-DDB for DNA. Binding of XPC-hHR23B to UV damaged DNA was very fast. Damaged DNA-XPC-hHR23B complexes were stable, with half of the complexes remaining four hours after challenge with excess W-damaged DNA at 30 degrees C. XPC-hHR23B had a higher level of affinity for (6-4) photoproducts than cyclobutane pyrimidine dimers, and some affinity for DNA treated with cisplatin and alkylating agents. XPC-hHR23B could bind to single-stranded M13 DNA, but only poorly to single-stranded homopolymers. The strong preference of XPC complex for structures in damaged duplex DNA indicates its importance as a primary damage recognition factor in non-transcribed DNA during human NER. (C) 2000 Academic Press.