Selby C P, Sancar A
Department of Biochemistry and Biophysics, University of North Carolina School of Medicine, Chapel Hill, North Carolina 27599, USA.
J Biol Chem. 1997 Jan 17;272(3):1885-90. doi: 10.1074/jbc.272.3.1885.
Transcription is coupled to repair in Escherichia coli and in humans. Proteins encoded by the mfd gene in E. coli and by the ERCC6/CSB gene in humans, both of which possess the so-called helicase motifs, are required for the coupling reaction. It has been shown that the Mfd protein is an ATPase but not a helicase and accomplishes coupling, in part, by disrupting the ternary complex of E. coli RNA polymerase stalled at the site of DNA damage. In this study we overproduced the human CSB protein using the baculovirus vector and purified and characterized the recombinant protein. CSB has an ATPase activity that is stimulated strongly by DNA; however, it neither acts as a helicase nor does it dissociate stalled RNA polymerase II, suggesting a coupling mechanism in humans different from that in prokaryotes. CSB is a DNA-binding protein, and it also binds to XPA, TFIIH, and the p34 subunit of TFIIE. These interactions are likely to play a role in recruiting repair proteins to ternary complexes formed at damage sites.
在大肠杆菌和人类中，转录与修复相偶联。大肠杆菌中由mfd基因编码的蛋白质以及人类中由ERCC6/CSB基因编码的蛋白质都参与了这种偶联反应，这两种蛋白质都具有所谓的解旋酶基序。研究表明，Mfd蛋白是一种ATP酶而非解旋酶，它通过破坏在DNA损伤位点停滞的大肠杆菌RNA聚合酶三元复合物来部分实现偶联。在本研究中，我们使用杆状病毒载体过量表达了人类CSB蛋白，并对重组蛋白进行了纯化和表征。CSB具有一种受DNA强烈刺激的ATP酶活性；然而，它既不充当解旋酶，也不解离停滞的RNA聚合酶II，这表明人类中的偶联机制与原核生物不同。CSB是一种DNA结合蛋白，它还能与XPA、TFIIH以及TFIIE的p34亚基结合。这些相互作用可能在将修复蛋白招募到损伤位点形成的三元复合物中发挥作用。
