Abstract
The variant form of the human syndrome xeroderma pigmentosum (XPV) is caused by a deficiency in DNA polymerase η (Polη), a DNA polymerase that enables replication through ultraviolet-induced pyrimidine dimers. Here we report high-resolution crystal structures of human Polη at four consecutive steps during DNA synthesis through cis-syn cyclobutane thymine dimers. Polη acts like a ‘molecular splint’ to stabilize damaged DNA in a normal B-form conformation. An enlarged active site accommodates the thymine dimer with excellent stereochemistry for two-metal ion catalysis. Two residues conserved among Polη orthologues form specific hydrogen bonds with the lesion and the incoming nucleotide to assist translesion synthesis. On the basis of the structures, eight Polη missense mutations causing XPV can be rationalized as undermining the molecular splint or perturbing the active-site alignment. The structures also provide an insight into the role of Polη in replicating through D loop and DNA fragile sites.
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Acknowledgements
We thank D. Leahy, M. Gellert and R. Craigie for critical reading of the manuscript. The research was funded by the intramural research program of NIDDK, NIH, and grants from the Ministry of Education, Culture, Sports, Science, and Technology of Japan. Y.Z. is the recipient of a Chinese Ministry of Education scholarship and joint PhD student in NIH-Zhejiang University Graduate Partnership Program. S.R.-M. received a fellowship from the Human Frontiers Science Program.
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C.B. determined the five structures; Y.Z. prepared the samples and grew the crystals; Y.K. did the kinetic and bypass assays; S.R.-M. determined the type 1 structure; M.G. prepared the clone and type 1 crystals; J.Y.L. made mutants; C.M. designed the functional assays; A.R.L. identified the unpublished XPV mutations; F.H. conceived the project; and W.Y. supervised the structure determination. C.B., Y.Z., F.H. and W.Y. prepared the manuscript. C.B. and Y.Z. contributed equally to the study. All authors discussed the results and commented on the manuscript.
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Biertümpfel, C., Zhao, Y., Kondo, Y. et al. Structure and mechanism of human DNA polymerase η. Nature 465, 1044–1048 (2010). https://doi.org/10.1038/nature09196
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DOI: https://doi.org/10.1038/nature09196
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