PMID:20852254

Citation	Lee, HW, Brice, AR, Wright, CB, Dominy, BN and Cao, W (2010) Identification of Escherichia coli mismatch-specific uracil DNA glycosylase as a robust xanthine DNA glycosylase. J. Biol. Chem. 285:41483-90
Abstract	The gene for the mismatch-specific uracil DNA glycosylase (MUG) was identified in the Escherichia coli genome as a sequence homolog of the human thymine DNA glycosylase with activity against mismatched uracil base pairs. Examination of cell extracts led us to detect a previously unknown xanthine DNA glycosylase (XDG) activity in E. coli. DNA glycosylase assays with purified enzymes indicated the novel XDG activity is attributable to MUG. Here, we report a biochemical characterization of xanthine DNA glycosylase activity in MUG. The wild type MUG possesses more robust activity against xanthine than uracil and is active against all xanthine-containing DNA (C/X, T/X, G/X, A/X and single-stranded X). Analysis of potentials of mean force indicates that the double-stranded xanthine base pairs have a relatively narrow energetic difference in base flipping, whereas the tendency for uracil base flipping follows the order of C/U > G/U > T/U > A/U. Site-directed mutagenesis performed on conserved motifs revealed that Asn-140 and Ser-23 are important determinants for XDG activity in E. coli MUG. Molecular modeling and molecular dynamics simulations reveal distinct hydrogen-bonding patterns in the active site of E. coli MUG that account for the specificity differences between E. coli MUG and human thymine DNA glycosylase as well as that between the wild type MUG and the Asn-140 and Ser-23 mutants. This study underscores the role of the favorable binding interactions in modulating the specificity of DNA glycosylases.
Links	PubMed PMC3009874 Online version:10.1074/jbc.M110.150003
Keywords	Amino Acid Sequence; Base Pair Mismatch; Catalytic Domain; DNA Glycosylases/chemistry; DNA Repair; Escherichia coli/metabolism; Humans; Hydrogen Bonding; Kinetics; Models, Molecular; Molecular Sequence Data; Mutagenesis, Site-Directed; Oligodeoxyribonucleotides/chemistry; Thymine/chemistry; Xanthine/chemistry
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Annotations

Gene product	Qualifier	GO Term	Evidence Code	with/from	Aspect	Extension	Notes	Status
ECOLI:MUG		GO:0004844 : uracil DNA N-glycosylase activity	ECO:0000314:		F		Figure 2A: Shows cleavage products of uracil-containing DNA substrates by MUG. Figure 3A: Shows kinetic analysis of MUG on uracil-containing substrates.	complete

References

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