Adenine excisional repair function of MYH protein on the adenine:8-hydroxyguanine base pair in double-stranded DNA

• DOI: 10.1093/nar/28.24.4912
• Publication Date: 2002-07-26T18:52:52Z
• Authors: K. Shinmura
• Keywords: 0301 basic medicine; Guanine; Base Sequence; DNA Repair; Base Pair Mismatch; Adenine; Escherichia coli Proteins; Carbon-Oxygen Lyases; Genetic Complementation Test; DNA; Deoxyribonuclease IV (Phage T4-Induced); DNA Glycosylases; Mitochondria; 3. Good health; Kinetics; 03 medical and health sciences; DNA-Formamidopyrimidine Glycosylase; Genes, Bacterial; Mutation; DNA-(Apurinic or Apyrimidinic Site) Lyase; Escherichia coli; Animals; Humans

Adenine paired with 8-hydroxyguanine (oh(8)G), a major component of oxidative DNA damage, is excised by MYH base excision repair protein in human cells. Since repair activity of MYH protein on an A:G mismatch has also been reported, we compared the repair activity of His(6)-tagged MYH proteins, expressed in Spodoptera frugiperda Sf21 cells, on A:oh(8)G and A:G mismatches by DNA cleavage assay and gel mobility shift assay. We also compared the repair ability of type 1 mitochondrial protein with type 2 nuclear protein, as well as of polymorphic type 1-Q(324) and 2-Q(310) proteins with type 1-H(324) and 2-H(310) proteins by DNA cleavage assay and complementation assay of an Escherichia coli mutM mutY strain. In a reaction buffer with a low salt (0-50 mM) concentration, adenine DNA glycosylase activity of type 2 protein was detected on both A:oh(8)G and A:G substrates. However, in a reaction buffer with a 150 mM salt concentration, similar to physiological conditions, the glycosylase activity on A:G, but not on A:oh(8)G, was extremely reduced and the binding activity of type 2 protein for A:G, but not for A:oh(8)G, was proportionally reduced. The glycosylase activity on A:oh(8)G and the ability to suppress spontaneous mutagenesis were greater for type 2 than type 1 enzyme. There was apparently no difference in the repair activities between the two types of polymorphic MYH proteins. These results indicate that human MYH protein specifically catalyzes the glycosylase reaction on A:oh(8)G under physiological salt concentrations.