Repair of thymine.guanine and uracil.guanine mismatched base-pairs in bacteriophage M13mp18 DNA heteroduplexes.
Repair of thymine.guanine (T.G) and uracil.guanine (U.G) mismatched base-pairs in bacteriophage M13mp18 replicative form (RF) DNA was compared upon transfection into repair-proficient or repair-deficient Escherichia coli strains. Oligonucleotide-directed mutagenesis was used to prepare covalently closed circular heteroduplexes that contained the mismatched base-pair at a restriction recognition site. The heteroduplexes were unmethylated at dam (5'-GATC-3') sites to avoid methylation-directed biasing of repair. In an E. coli host containing uracil-DNA glycosylase (ung+), about 97% of the transfecting U.G-containing heteroduplexes had the U residue excised by the uracil-excision repair system. With the analogous T.G mispair, mismatch repair operated on almost all of the transfecting heteroduplexes and removed the T residue in about 75% of them when the mismatched T was on the minus strand of the RF DNA. Similar preferential excision of the minus-strand's mismatched base was observed whether the heteroduplex RF DNA molecules had only one or both strands unmethylated at dcm (5'-CC(A/T)GG-3') sites and whether the RF DNA was prepared by primer extension in vitro or by reannealing mutant and non-mutant DNA strands. Also, the extent and directionality of repair was the same at a U.G mispair in ung- host cells as at the analogous T.G mispair in ung- or ung+ cells. Only in a mismatch repair-deficient (mutH-) host was the plus strand of the transfecting M13mp18 heteroduplex DNA preferentially repaired. It is suggested that the plus strand nick made by the M13-encoded gene II protein might be employed by a mutH- host to initiate repair on that strand.
Shenoy
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Department of Biochemistry, Tulane Medical School, New Orleans, LA 70112.
Ehrlich
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Ehrlich
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D001435:Bacteriophages / Q000235:genetics*; D001482:Base Composition; D004260:DNA Repair; D004279:DNA, Viral; D004926:Escherichia coli / Q000235:genetics; D006147:Guanine; D008745:Methylation; D009692:Nucleic Acid Heteroduplexes; D013941:Thymine; D014498:Uracil
D004279:DNA, Viral; D009692:Nucleic Acid Heteroduplexes; D014498:Uracil; D006147:Guanine; D013941:Thymine