Constantinescu-Aruxandei Diana, Petrovic-Stojanovska Biljana, Penedo J Carlos, White Malcolm F, Naismith James H
Biomedical Sciences Research Complex, University of St Andrews, Fife KY16 9ST, UK.
Biomedical Sciences Research Complex, University of St Andrews, Fife KY16 9ST, UK
Nucleic Acids Res. 2016 Apr 7;44(6):2806-15. doi: 10.1093/nar/gkw102. Epub 2016 Feb 20.
The xeroderma pigmentosum group D (XPD) helicase is a component of the transcription factor IIH complex in eukaryotes and plays an essential role in DNA repair in the nucleotide excision repair pathway. XPD is a 5' to 3' helicase with an essential iron-sulfur cluster. Structural and biochemical studies of the monomeric archaeal XPD homologues have aided a mechanistic understanding of this important class of helicase, but several important questions remain open. In particular, the mechanism for DNA loading, which is assumed to require large protein conformational change, is not fully understood. Here, DNA binding by the archaeal XPD helicase from Thermoplasma acidophilum has been investigated using a combination of crystallography, cross-linking, modified substrates and biochemical assays. The data are consistent with an initial tight binding of ssDNA to helicase domain 2, followed by transient opening of the interface between the Arch and 4FeS domains, allowing access to a second binding site on helicase domain 1 that directs DNA through the pore. A crystal structure of XPD from Sulfolobus acidocaldiarius that lacks helicase domain 2 has an otherwise unperturbed structure, emphasizing the stability of the interface between the Arch and 4FeS domains in XPD.
着色性干皮病D组（XPD）解旋酶是真核生物中转录因子IIH复合物的一个组成部分，在核苷酸切除修复途径的DNA修复中起关键作用。XPD是一种具有必需铁硫簇的5'至3'解旋酶。对单体古细菌XPD同源物的结构和生化研究有助于对这一重要解旋酶类别进行机制理解，但仍有几个重要问题悬而未决。特别是，DNA加载机制（假定需要大的蛋白质构象变化）尚未完全理解。在此，利用晶体学、交联、修饰底物和生化分析相结合的方法，研究了嗜热栖热菌的古细菌XPD解旋酶与DNA的结合。数据表明，单链DNA最初紧密结合到解旋酶结构域2，随后Arch和4FeS结构域之间的界面短暂打开，从而可以进入解旋酶结构域1上的第二个结合位点，引导DNA穿过孔隙。来自嗜酸嗜热栖热菌的缺乏解旋酶结构域2的XPD晶体结构具有未受干扰的结构，强调了XPD中Arch和4FeS结构域之间界面的稳定性。
