DNA polymerase I (Klenow fragment): Role of the structure and length of a template in enzyme recognition

• Published in: FEBS letters Vol. 248; no. 1-2; pp. 97 - 100
• Main Authors: Kolocheva, T.I., Nevinsky, G.A., Volchkova, V.A., Levina, A.S., Khomov, V.V., Lavrik, O.I.
• Format: Journal Article
• Language: English
• Published: Amsterdam Elsevier B.V 08.05.1989; Elsevier
• Subjects: Analytical, structural and metabolic biochemistry; Biological and medical sciences; BSA, bovine serum albumin; DNA; DNA Polymerase I; DNA-Directed DNA Polymerase; Electrochemistry; Enzymes and enzyme inhibitors; Fundamental and applied biological sciences. Psychology; Hydrogen Bonding; Kinetics; Klenow fragment; KLF, Klenow fragment; Models, Theoretical; Molecular Structure; Oligonucleotides; Peptide Fragments; Protein-nucleic acid interaction; R1, d(Tp)2C[Pt2+-(NH3)2OH](pT)7; R2, d[(Tp)2C(Pt2(NH3)2OH)p]3T; Templates, Genetic; Transferases; BSA, bovine serum albumin; Klenow fragment; R1, d(Tp)2C[Pt2+-(NH3)2OH](pT)7; Protein-nucleic acid interaction; R2, d[(Tp)2C(Pt2(NH3)2OH)p]3T; KLF, Klenow fragment; DNA polymerase I; Human; Phosphates; Enzyme; Electrostatic interaction; Hydrophobic interaction; Dissociation constant; Inactivation; Ion exchange chromatography; Enzymatic activity; Structure activity relation; DNA polymerase 1; Molecular association; Models; Oligonucleotide; Hydrogen bond
• ISSN: 0014-5793; 1873-3468
• DOI: 10.1016/0014-5793(89)80439-9

The values of Kd and Gibbs energy (ΔG°) have been measured for complexes of the template site of DNA polymerase I Klenow fragment with the homo-oligonucleotides d(pC)n, d(pT)n, d(pG)n and d(pA)n and hetero-oligonucleotides of various structures and lengths. These parameters were evaluated from the protective effect of the oligonucleotide on enzyme inactivation by the affinity reagents d(Tp)2C[Pt2+(NH3)2OH](pT)7 and d[(Tp)2C(Pt2+(NH3)2OH)p]3T of the template site. The present results and previously reported data [(1985) Biorg. Khim. 13, 357–369] indicate that the nucleoside components of the template form complexes as a result of their hydrophobic interactions with the enzyme. Only one template internucleotide phosphate forms an Me2+-dependent electrostatic contact and a hydrogen bond with the enzyme. The 19–20-nucleotide fragments of the template appear to interact with the protein molecule.