PMID:10924147
| Citation |
Kamiya, H, Maki, H and Kasai, H (2000) Two DNA polymerases of Escherichia coli display distinct misinsertion specificities for 2-hydroxy-dATP during DNA synthesis. Biochemistry 39:9508-13 |
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| Abstract |
The insertion specificities of an oxidized dATP analogue, 2-hydroxydeoxyadenosine 5'-triphosphate (2-OH-dATP), were determined using the alpha (catalytic) subunit of Escherichia coli DNA polymerase III and the exonuclease-deficient Klenow fragment of DNA polymerase I. In contrast to our previous observation that mammalian DNA polymerase alpha incorporated the oxidized nucleotide opposite T and C, these two E. coli DNA polymerases incorporated 2-OH-dATP opposite T and G on the DNA template. Steady-state kinetic studies indicated that the alpha subunit incorporated 2-OH-dATP 10 times more frequently opposite T than opposite G. On the other hand, the incorporation of 2-OH-dATP opposite T by the exonuclease-deficient Klenow fragment was 2 orders of magnitude more efficient than that opposite G. These results indicate that the misinsertion specificity of 2-OH-dATP differs between replicative and repair-type DNA polymerases, and provide a biochemical basis for the mutations induced by 2-OH-dATP in E. coli. |
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| Keywords |
Adenosine Triphosphate/analogs & derivatives; Adenosine Triphosphate/metabolism; Catalysis; DNA Polymerase I/genetics; DNA Polymerase I/metabolism; DNA Polymerase III/metabolism; DNA Replication; Escherichia coli/enzymology; Escherichia coli/genetics; Exodeoxyribonucleases/deficiency; Exodeoxyribonucleases/genetics; Kinetics; Reactive Oxygen Species/metabolism; Substrate Specificity/genetics |
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