Strand-specific recognition of DNA damages by XPD provides insights into Nucleotide excision repair substrate versatility

Abstract

Background: XPD is important for DNA lesion recognition by the nucleotide excision repair (NER) system. Results: Dependent on the lesion type, XPD recognizes lesions either on the protein-translocated or on the nontranslocated DNA strand. Conclusion: XPD employs different recognition strategies for different types of damage. Significance: Different lesion-specific recognition approaches may enhance the remarkably broad target spectrum of NER. Recognition and removal ofDNAdamages is essential for cellular and organismal viability. Nucleotide excision repair (NER) is the sole mechanism in humans for the repair of carcinogenic UV irradiation-induced photoproducts in the DNA, such as cyclobutane pyrimidine dimers. The broad substrate versatility of NER further includes, among others, various bulky DNA adducts. It has been proposed that the 5'-3' helicase XPD (xeroderma pigmentosum group D) protein plays a decisive role in damage verification. However, despite recent advances such as the identification of a DNA-binding channel and central pore in the protein, through which the DNA is threaded, as well as a dedicated lesion recognition pocket near the pore, the exact process of target site recognition and verification in eukaryotic NER still remained elusive. Our single molecule analysis by atomic force microscopy reveals for the first time that XPD utilizes different recognition strategies to verify structurally diverse lesions. Bulky fluorescein damage is preferentially detected on the translocated strand, whereas the opposite strand preference is observed for a cyclobutane pyrimidine dimer lesion. Both states, however, lead to similar conformational changes in the resulting specific complexes, indicating a merge to a "final" verification state, which may then trigger the recruitment of further NER proteins.© 2014 by The American Society for Biochemistry and Molecular Biology, Inc.