The affinity of the Klenow fragment of E. coli DNA-polymerase 1 to primers containing bases noncomplementary to the template and hairpin-like elements.

Abstract:

The Km and Vmax values for a set of primers: d(pT)n (pC) (pT)m (n = 3-9, m = 0-7) and d(pT)4 (pCpG)k (pT)4 (k = 1-5) have been estimated. Poly(dA) was used as a template. The number of complementary bases from the 3' end to a noncomplementary ones was shown to determine the efficiency of interaction of d(pT)n (pC) (pT)m with the Klenow fragment. Oligonucleotides d(pT)4 (pCpG)k (pT)4, in solution forming duplexes containing hairpin-like elements, show a higher affinity to the enzyme than control d(pT)4, d(pT)8 and d(pT)n (pC) (pT)m primers. For example, the Km value (1.1 nM) for d(pT)4 (pCpG)5 (pT)4 is about 14,000 and 200 times lower than those for d(pT)4 and d(pT)8, respectively. Possible reasons for such an abnormally high affinity of the above primers are discussed.

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