Molecular cloning and functional analysis of the MutY homolog of Deinococcus radiodurans.
The mutY homolog gene (mutY(Dr)) from Deinococcus radiodurans encodes a 39.4-kDa protein consisting of 363 amino acids that displays 35% identity to the Escherichia coli MutY (MutY(Ec)) protein. Expressed MutY(Dr) is able to complement E. coli mutY mutants but not mutM mutants to reduce the mutation frequency. The glycosylase and binding activities of MutY(Dr) with an A/G-containing substrate are more sensitive to high salt and EDTA concentrations than the activities with an A/7,8-dihydro-8-oxoguanine (GO)-containing substrate are. Like the MutY(Ec) protein, purified recombinant MutY(Dr) expressed in E. coli has adenine glycosylase activity with A/G, A/C, and A/GO mismatches and weak guanine glycosylase activity with a G/GO mismatch. However, MutY(Dr) exhibits limited apurinic/apyrimidinic lyase activity and can form only weak covalent protein-DNA complexes in the presence of sodium borohydride. This may be due to an arginine residue that is present in MutY(Dr) at the position corresponding to the position of MutY(Ec) Lys142, which forms the Schiff base with DNA. The kinetic parameters of MutY(Dr) are similar to those of MutY(Ec). Although MutY(Dr) has similar substrate specificity and a binding preference for an A/GO mismatch over an A/G mismatch, as MutY(Ec) does, the binding affinities for both mismatches are slightly lower for MutY(Dr) than for MutY(Ec). Thus, MutY(Dr) can protect the cell from GO mutational effects caused by ionizing radiation and oxidative stress.
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Department of Biochemistry and Molecular Biology, School of Medicine, University of Maryland, Baltimore, Maryland 21201, USA.
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D000225:Adenine / Q000378:metabolism; D000595:Amino Acid Sequence; D001426:Bacterial Proteins; D020137:Base Pair Mismatch; D019757:Carbon-Oxygen Lyases / Q000378:metabolism; D003001:Cloning, Molecular; D045647:DNA Glycosylases; D004260:DNA Repair; D043603:DNA-(Apurinic or Apyrimidinic Site) Lyase; D043223:Deoxyribonuclease IV (Phage T4-Induced); D004492:Edetic Acid / Q000494:pharmacology; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D029968:Escherichia coli Proteins; D005816:Genetic Complementation Test; D006147:Guanine / Q000378:metabolism; D007700:Kinetics; D008969:Molecular Sequence Data; D009154:Mutation; D009699:N-Glycosyl Hydrolases / Q000235:genetics* / Q000502:physiology*; D017386:Sequence Homology, Amino Acid; D012965:Sodium Chloride / Q000494:pharmacology; D013824:Thermus / Q000201:enzymology* / Q000235:genetics*
D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; D012965:Sodium Chloride; D006147:Guanine; D004492:Edetic Acid; D043223:Deoxyribonuclease IV (Phage T4-Induced); C468972:endonuclease IV, E coli; D045647:DNA Glycosylases; C439046:MutY protein, Deinococcus radiodurans; D009699:N-Glycosyl Hydrolases; C061738:mutY adenine glycosylase; D019757:Carbon-Oxygen Lyases; D043603:DNA-(Apurinic or Apyrimidinic Site) Lyase; D000225:Adenine