DNA polymerase accuracy and spontaneous mutation rates: frequencies of purine.purine, purine.pyrimidine, and pyrimidine.pyrimidine mismatches during DNA replication.

Abstract:

DNA from the am16 mutant of bacteriophage phi X174 may be replicated in vitro and expressed in vivo to give five classes of revertants. Each class may be specifically induced by the appropriate biasing of the concentrations of deoxynucleoside triphosphates in a predictable manner. The frequency of each reversion follows a kinetic rate equation relating it to the concentrations of the triphosphates involved in the substitution. The reversions corresponding to TAG leads to GAG, AAG, CAG, TGG, and TCG are calculated to occur with frequencies of 5 X 10(-7), 4 X 10(-7), 4 X 10(-7), approximately 2 X 10(-7), and approximately 5 X 10(-9), respectively, at the concentration of triphosphates found in vivo. The frequencies are in the range found for the reversion of the phage in vivo and so are consistent with errors in nucleotide selection by DNA polymerase (deoxynucleosidetriphosphate:DNA deoxynucleotidyltransferase, EC 2.7.7.7) III being largely responsible for the rate of spontaneous mutation in vivo. The relative frequency of mispairing leading to misincorporation is: purine.purine approximately purine.pyrimidine much greater than pyrimidine.pyrimidine, confirming predictions from model-building studies that transversions arise through purine.purine mismatches.

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