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The affinity of the Klenow fragment of E. coli DNA‐polymerase 1 to primers containing bases noncomplementary to the template and hairpin‐like elements
Author(s) -
Ljach Marija V.,
Kolocheva Tatjana I.,
Gorn Vladimir V.,
Levina Asja S.,
Nevinsky Georgy A.
Publication year - 1992
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/0014-5793(92)80155-a
Subject(s) - klenow fragment , dna polymerase i , chemistry , oligonucleotide , stereochemistry , microbiology and biotechnology , dna polymerase , dna , crystallography , polymerase chain reaction , biology , exonuclease , biochemistry , reverse transcriptase , gene
The K m and V max values for a set of primers: d(pT) n (pC)(pT) m ( n =3–9. m =0–7) and d(pT) 4 (pCpG) 4 (pT) 4 ( k =1–5) have been estimated. Poly(dA) was used as template. The number of complementary bases from the 3′ end to a noncomplementatry 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) 4 (pT) 4 in solution forming duplexes containing hairpin‐like elements, show a higher affinity to the enzyme that control d(pT) 4 , d(pT) n , and d(pT) n (pC)(pT) m primers. For example, the K m 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) ? respectively. Possible reasons for such an abnormally high affinity of the above primers are discussed.