Intact MutY and its catalytic domain differentially contact with A/8-oxoG-containing DNA.
Escherichia coli MutY is an adenine and a weak guanine DNA glycosylase active on DNA substrates containing A/G, A/8-oxoG, A/C or G/8-oxoG mismatches. A truncated form of MutY (M25, residues 1-226) retains catalytic activity; however, the C-terminal domain of MutY is required for specific binding to the 8-oxoG and is critical for mutation avoidance of oxidative damage. Using alkylation interference experiments, the determinants of the truncated and intact MutY were compared on A/8-oxoG-containing DNA. Several purines within the proximity of mismatched A/8-oxoG show differential contact by the truncated and intact MutY. Most importantly, methylation at the N7 position of the mismatched 8-oxoG and the N3 position of mismatched A interfere with intact MutY but not with M25 binding. The electrostatic contacts of MutY and M25 with the A/8-oxoG-containing DNA substrates are drastically different as shown by ethylation interference experiments. Five consecutive phosphate groups surrounding the 8-oxoG (one on the 3' side and four on the 5' side) interact with MutY but not with M25. The activities of the truncated and intact MutY are modulated differently by two minor groove-binding drugs, distamycin A and Hoechst 33258. Both distamycin A and Hoechst 33258 can inhibit, to a similar extent, the binding and glycosylase activities of MutY and M25 on A/G mismatch. However, binding and glycosylase activities on A/8-oxoG mismatch of intact MutY are inhibited to a lesser degree than those of M25. Overall, these results suggest that the C-terminal domain of MutY specifies additional contact sites on A/GO-containing DNA that are not found in MutY-A/G and M25-A/8-oxoG interactions.
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Department of Biochemistry and Molecular Biology, School of Medicine, University of Maryland, Baltimore, MD 21201, USA.
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D000478:Alkylation; D001483:Base Sequence; D006690:Bisbenzimidazole / Q000494:pharmacology; D020134:Catalytic Domain; D004247:DNA / Q000737:chemistry / Q000187:drug effects / Q000378:metabolism; D004249:DNA Damage; D045647:DNA Glycosylases; D019175:DNA Methylation; D004260:DNA Repair; D004214:Distamycins / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D006147:Guanine / Q000031:analogs & derivatives* / Q000737:chemistry / Q000378:metabolism*; D009154:Mutation; D009699:N-Glycosyl Hydrolases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D009690:Nucleic Acid Conformation; D011485:Protein Binding / Q000187:drug effects
D004214:Distamycins; C024829:8-hydroxyguanine; D006147:Guanine; C030000:stallimycin; D004247:DNA; D045647:DNA Glycosylases; D009699:N-Glycosyl Hydrolases; C061738:mutY adenine glycosylase; D006690:Bisbenzimidazole