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The algorithm of estimation of theKmvalues for primers of various structure and length in the polymerization reaction catalyzed by Klenow fragment of DNA polymerase I fromE.coli

Authors
Journal
FEBS Letters
0014-5793
Publisher
Wiley Blackwell (John Wiley & Sons)
Publication Date
Volume
258
Issue
1
Identifiers
DOI: 10.1016/0014-5793(89)81642-4
Keywords
  • Klenow Fragment
  • Primer
  • Kmcalculation
  • Algorithm Of
Disciplines
  • Biology

Abstract

Abstract DNA synthesis at primers d(pT) n , d(pA) n , d(pC) n , and d(pG) n in the presence of corresponding complementary templates and at hetero-oligoprimers complementary to MB phage DNA was investigated. The values of both -log K m and log V max increased linearly if homo-oligoprimers contained less than 10 nucleotides. The lengthening of d(pT) n and d(pA) n primers by one mononucleotide unit ( n = 1–10) resulted in the 1.82-fold decrease of the K m values. The incremental decreases of K m for d(pC) n and d(pG) n were equal to about 2.46. The enhancement of the homo- and heterooligonucleotide primers' affinity to the enzyme due to one Watson-Crick hydrogen bond between complementary template and primer is about 1.35 times. This allows to calculate the K m values for primers of various structure and length up to 10 units. The objective laws of the K m and V max values changes for primers containing more than 10 nucleotides were analyzed.

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