Base incorporation and extension at a site-specific ethenocytosine by Escherichia coli DNA polymerase I Klenow fragment.
Ethenocytosine (epsilon C) is a highly mutagenic exocyclic DNA lesion induced by carcinogens vinyl chloride and urethane. We have examined base incorporation and extension at a site-specific epsilon C residue by a quantitative gel electrophoretic assay using an exonuclease-deficient version of Escherichia coli DNA polymerase I (Klenow fragment) as the model enzyme. The data show that the KM for incorporation of adenine or thymine opposite epsilon C by is about 5 orders of magnitude higher than that for the incorporation of guanine opposite normal cytosine. The KM for base extension past epsilon C:A and epsilon C:T pairs is 1-2 orders of magnitude higher than that observed for a C:G pair. Although adenine misinsertion is favored over that of thymine, base extension occurs more readily when the base incorporated opposite epsilon C is thymine.
Simha
D
D
Department of Microbiology and Molecular Genetics, UMD-New Jersey Medical School, Newark 07103-2714.
Yadav
D
D
Rzepka
R W
RW
Palejwala
V A
VA
Humayun
M Z
MZ
D000477:Alkylating Agents / Q000378:metabolism / Q000633:toxicity; D001482:Base Composition; D001483:Base Sequence; D003596:Cytosine / Q000031:analogs & derivatives* / Q000737:chemistry / Q000378:metabolism / Q000633:toxicity; D004247:DNA / Q000737:chemistry / Q000378:metabolism*; D004249:DNA Damage; D004256:DNA Polymerase I / Q000235:genetics* / Q000378:metabolism; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009153:Mutagens / Q000378:metabolism* / Q000633:toxicity; D009838:Oligodeoxyribonucleotides / Q000378:metabolism; D013698:Templates, Genetic
C018289:3,N(4)-ethenocytosine; D000477:Alkylating Agents; D009153:Mutagens; D009838:Oligodeoxyribonucleotides; D003596:Cytosine; D004247:DNA; D004256:DNA Polymerase I