Characterization of the recombinant MutY homolog, an adenine DNA glycosylase, from yeast Schizosaccharomyces pombe.
The mutY homolog (SpMYH) gene from a cDNA library of Schizosaccharomyces pombe encodes a protein of 461 amino acids that displays 28 and 31% identity to Escherichia coli MutY and human MutY homolog (MYH), respectively. Expressed SpMYH is able to complement an E. coli mutY mutant to reduce the mutation rate. Similar to E. coli MutY protein, purified recombinant SpMYH expressed in E. coli has adenine DNA glycosylase and apurinic/apyrimidinic lyase activities on A/G- and A/7,8-dihydro-8-oxoguanine (8-oxoG)-containing DNA. However, both enzymes have different salt requirements and slightly different substrate specificities. SpMYH has greater glycosylase activity on 2-aminopurine/G and A/2-aminopurine but weaker activity on A/C than E. coli MutY. Both enzymes also have different substrate binding affinity and catalytic parameters. Although SpMYH has great affinity to A/8-oxoG-containing DNA as MutY, the binding affinity to A/G-containing DNA is substantially lower for SpMYH than MutY. SpMYH has similar reactivity to both A/G- and A/8-oxoG-containing DNA; however, MutY cleaves A/G-containing DNA about 3-fold more efficiently than it does A/8-oxoG-containing DNA. Thus, SpMYH is the functional eukaryotic MutY homolog responsible for reduction of 8-oxoG mutational effect.
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Department of Biochemistry and Molecular Biology, University of Maryland, Baltimore, Maryland 21201, USA. aluchang@umaryland.edu
Fawcett
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D000595:Amino Acid Sequence; D001483:Base Sequence; D045647:DNA Glycosylases; D017931:DNA Primers; D004492:Edetic Acid; D004926:Escherichia coli / Q000235:genetics; D007700:Kinetics; D008969:Molecular Sequence Data; D009154:Mutation; D009699:N-Glycosyl Hydrolases / Q000737:chemistry / Q000378:metabolism*; D009692:Nucleic Acid Heteroduplexes; D011485:Protein Binding; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D012492:Salts; D012568:Schizosaccharomyces / Q000201:enzymology*; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity
D017931:DNA Primers; D009692:Nucleic Acid Heteroduplexes; D011994:Recombinant Proteins; D012492:Salts; D004492:Edetic Acid; D045647:DNA Glycosylases; D009699:N-Glycosyl Hydrolases; C061738:mutY adenine glycosylase