title: Effect of point substitutions within the minimal DNA-binding domain of xeroderma pigmentosum group A protein on interaction with DNA intermediates of nucleotide excision repair.
abstract: Xeroderma pigmentosum factor A (XPA) is one of the key proteins in the nucleotide excision repair (NER) process. The effects of point substitutions in the DNA-binding domain of XPA (positively charged lysine residues replaced by negatively charged glutamate residues: XPA K204E, K179E, K141E, and tandem mutant K141E/K179E) on the interaction of the protein with DNA structures modeling intermediates of the damage recognition and pre-incision stages in NER were analyzed. All these mutations decreased the affinity of the protein to DNA, the effect depending on the substitution and the DNA structure. The mutant as well as wild-type proteins bind with highest efficiency partly open damaged DNA duplex, and the affinity of the mutants to this DNA is reduced in the order: K204E > K179E >> K141E = K141/179E. For all the mutants, decrease in DNA binding efficiency was more pronounced in the case of full duplex and single-stranded DNA than with bubble-DNA structure, the difference between protein affinities to different DNA structures increasing as DNA binding activity of the mutant decreased. No effect of the studied XPA mutations on the location of the protein on the partially open DNA duplex was observed using photoinduced crosslinking with 5-I-dUMP in different positions of the damaged DNA strand. These results combined with earlier published data suggest no direct correlation between DNA binding and activity in NER for these XPA mutants.
authors.affiliation: Institute of Chemical Biology and Fundamental Medicine, Siberian Branch of the Russian Academy of Sciences, Novosibirsk, 630090, Russia. nadyarec@niboch.nsc.ru.
authors.forename: E A
authors.initials: EA
authors.lastname: Maltseva
authors.forename: Y S
authors.initials: YS
authors.lastname: Krasikova
authors.forename: H
authors.initials: H
authors.lastname: Naegeli
authors.forename: O I
authors.initials: OI
authors.lastname: Lavrik
authors.forename: N I
authors.initials: NI
authors.lastname: Rechkunova
mesh_terms: D019943:Amino Acid Substitution; D001483:Base Sequence; D004247:DNA / Q000737:chemistry / Q000378:metabolism*; D004249:DNA Damage; D004260:DNA Repair; D004926:Escherichia coli / Q000378:metabolism; D006801:Humans; D009690:Nucleic Acid Conformation; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics; D051760:Xeroderma Pigmentosum Group A Protein / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
chemical_list: D011994:Recombinant Proteins; D051760:Xeroderma Pigmentosum Group A Protein; D004247:DNA