The elongation of mismatched primers by DNA polymerase alpha from calf thymus.
The ability of the 9S and 5.7S DNA polymerase alpha subspecies from calf thymus in elongating a mismatched primer terminus has been investigated. With poly(dA) as template, the elongation rate for (dT)8dG, (dT)8dC and (dT)10dGdT is 20-fold lower for the 9S enzyme and 5-fold lower for the 5.7S enzyme as compared to (dT)10. The presence of a second mismatch at the primer terminus reduces the elongation rate further by a factor of two. Exonucleolytic excision of the mismatches can be excluded. With (dT)8dG (dT)n as primer we show, that at least five T-residues have to follow the mismatch in order to establish the elongation rate of a perfectly paired primer. The KM value for (dT)10 dG as primer is 400 nM as compared to 10 nM for (dT)10. Addition of Mn2+ increases the relative efficiency of elongation of the mismatched primers.
D000818:Animals; D002417:Cattle; D004257:DNA Polymerase II / Q000378:metabolism*; D007700:Kinetics; D008274:Magnesium / Q000494:pharmacology; D008345:Manganese / Q000494:pharmacology; D013329:Structure-Activity Relationship; D013379:Substrate Specificity; D013698:Templates, Genetic; D013950:Thymus Gland / Q000201:enzymology*
D008345:Manganese; D004257:DNA Polymerase II; D008274:Magnesium