2Novosibirsk State University, ul. Pirogova 2, 630090 Novosibirsk, Russia
3Faculty of Chemistry, Lomonosov Moscow State University, 119992 Moscow, Russia
4Department of Pharmacological Sciences, SUNY Stony Brook, Stony Brook, New York 11794-3400, USA; E-mail: Orlando@pharm.stonybrook.edu
* To whom correspondence should be addressed.
Received December 20, 2007; Revision received March 18, 2008
The interaction of nucleotide excision repair factors--xeroderma pigmentosum complementation group C protein in complex with human homolog of yeast Rad23 protein (XPC-HR23B), replication protein A (RPA), and xeroderma pigmentosum complementation group A protein (XPA)--with 48-mer DNA duplexes imitating damaged DNA structures was investigated. All studied proteins demonstrated low specificity in binding to damaged DNA compared with undamaged DNA duplexes. RPA stimulates formation of XPC-HR23B complex with DNA, and when XPA and XPC-HR23B are simultaneously present in the reaction mixture a synergistic effect in binding of these proteins to DNA is observed. RPA crosslinks to DNA bearing photoreactive 5I-dUMP residue on one strand and fluorescein-substituted dUMP analog as a lesion in the opposite strand of DNA duplex and also stimulates cross-linking with XPC-HR23B. Therefore, RPA might be one of the main regulation factors at various stages of nucleotide excision repair. The data are in agreement with the cooperative binding model of nucleotide excision repair factors participating in pre-incision complex formation with DNA duplexes bearing damages.
KEY WORDS: nucleotide excision repair factors, photoaffinity labeling, photoreactive oligonucleotides, complex formation