Recognition of nonhybridizing base pairs during nucleotide excision repairof DNA

Nondistorting C4' backbone adducts serve as molecular tools to analyze the strategy by which a limited number of human nucleotide excision repair (NER ) factors recognize an infinite variety of DNA lesions. We have constructed composite DNA substrates containing a noncomplementary site adjacent to a nondistorting C4' adduct to show that the loss of hydrogen bonding contacts between partner strands is an essential signal for the recruitment of NER enzymes. This specific conformational requirement for excision is mediated by the affinity of xeroderma pigmentosum group A (XPA) protein for nonhybri dizing sites in duplex DNA. XPA recognizes defective Watson-Crick base pair conformations even in the absence of DNA adducts or other covalent modific ations, apparently through detection of hydrophobic base components that ar e abnormally exposed to the double helical surface. This recognition functi on of XPA is enhanced by replication protein A (RPA) such that, in combinat ion, XPA and RPA constitute a potent molecular sensor of denatured base pai rs. Our results indicate that the XPA-RPA complex may promote damage recogn ition by monitoring Watson-Crick base pair integrity, thereby recruiting th e human NER system preferentially to sites where hybridization between comp lementary strands is weakened or entirely disrupted.