title: A mutant HpaII methyltransferase functions as a mutator enzyme.
abstract: DNA (cytosine-5)-methyltransferases can cause deamination of cytosine when the cofactor S-adenosylmethionine (AdoMet) is limiting and thus function as sequence-specific C-->U mutator enzymes. Here we explored whether mutations causing inactivation of the cofactor binding activity of the HpaII methyltransferase, thus mimicking conditions of limiting AdoMet concentration, could convert a DNA methyltransferase to a C-->U mutator enzyme. We created two mutator enzymes from the HpaII methyltransferase (F38S and G40D) which both showed enhanced cytosine deamination activities in vitro and in vivo. Interestingly, the G:U mispairs generated by these enzymes were not repaired completely in bacteria equipped with uracil-DNA glycosylase-initiated repair machinery, giving rise to a potent mutator phenotype. This is the first report showing the creation of mutator enzymes from a DNA methyltransferase and the demonstration of their mutagenicity in living cells.
authors.affiliation: Department of Biochemistry and Molecular Biology, USC/Norris Comprehensive Cancer Center, University of Southern California, Los Angeles 90033, USA.
authors.forename: J C
authors.initials: JC
authors.lastname: Shen
authors.forename: J M
authors.initials: JM
authors.lastname: Zingg
authors.forename: A S
authors.initials: AS
authors.lastname: Yang
authors.forename: C
authors.initials: C
authors.lastname: Schmutte
authors.forename: P A
authors.initials: PA
authors.lastname: Jones
mesh_terms: D001483:Base Sequence; D003596:Cytosine / Q000378:metabolism*; D004247:DNA / Q000378:metabolism; D015257:DNA-Cytosine Methylases / Q000235:genetics* / Q000378:metabolism; D003641:Deamination; D003841:Deoxycytidine / Q000031:analogs & derivatives / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D006190:Haemophilus / Q000201:enzymology* / Q000235:genetics; D008745:Methylation; D008969:Molecular Sequence Data; D016296:Mutagenesis; D016297:Mutagenesis, Site-Directed; D016133:Polymerase Chain Reaction; D011994:Recombinant Proteins / Q000378:metabolism
chemical_list: D011994:Recombinant Proteins; D003841:Deoxycytidine; D003596:Cytosine; D004247:DNA; C029641:DNA modification methylase HpaII; D015257:DNA-Cytosine Methylases; C007746:5-fluoro-2'-deoxycytidine