title: Comparative analysis of interaction of human and yeast DNA damage recognition complexes with damaged DNA in nucleotide excision repair.
abstract: The human XPC-RAD23B complex and its yeast ortholog, Rad4-Rad23, are the primary initiators of global genome nucleotide excision repair. The interaction of these proteins with damaged DNA was analyzed using model DNA duplexes containing a single fluorescein-substituted dUMP analog as a lesion. An electrophoretic mobility shift assay revealed similarity between human and yeast proteins in DNA binding. Quantitative analyses of XPC/Rad4 binding to the model DNA structures were performed by fluorescent depolarization measurements. XPC-RAD23B and Rad4-Rad23 proteins demonstrate approximately equal binding affinity to the damaged DNA duplex (K(D) ∼ (0.5 ± 0.1) and (0.6 ± 0.3) nM, respectively). Using photoreactive DNA containing 5-iodo-dUMP in defined positions, XPC/Rad4 location on damaged DNA was shown. Under conditions of equimolar binding to DNA both proteins exhibited the highest level of cross-links to 5I-dUMP located exactly opposite the damaged nucleotide. The positioning of the XPC and Rad4 proteins on damaged DNA by photocross-linking footprinting is consistent with x-ray analysis of the Rad4-DNA crystal complex. The identity of the XPC and Rad4 location illustrates the common principles of structure organization of DNA damage-scanning proteins from different Eukarya organisms.
authors: Institute of Chemical Biology and Fundamental Medicine, Novosibirsk 630090, Russia.
authors: Yuliya S
authors: YS
authors: Krasikova
authors: Nadejda I
authors: NI
authors: Rechkunova
authors: Ekaterina A
authors: EA
authors: Maltseva
authors: Pavel E
authors: PE
authors: Pestryakov
authors: Irina O
authors: IO
authors: Petruseva
authors: Kaoru
authors: K
authors: Sugasawa
authors: Xuejing
authors: X
authors: Chen
authors: Jung-Hyun
authors: JH
authors: Min
authors: Olga I
authors: OI
authors: Lavrik
mesh_terms: D004249:DNA Damage / Q000502:physiology*; D004260:DNA Repair / Q000502:physiology*; D045643:DNA Repair Enzymes / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D046912:Multiprotein Complexes / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
chemical_list: D004268:DNA-Binding Proteins; D046912:Multiprotein Complexes; D029701:Saccharomyces cerevisiae Proteins; D045643:DNA Repair Enzymes