Pyridoxal 5'-phosphate mediated inactivation of Escherichia coli DNA polymerase I: identification of lysine-635 as an essential residue for the processive mode of DNA synthesis

Inactivation of Escherichia coli DNA polymerase I by pyridoxal 5'-phosphate treatment results from its reactivity at multiple lysine residues. One of these residues, lysine-758, has been shown to be located at the substrate binding site in DNA polymerase I [Basu, A., & Modak, M. J. (1987) B...

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Bibliographic Details
Published in:Biochemistry (Easton) 1988-09, Vol.27 (18), p.6710-6716
Main Authors: Basu, Subhalakshmi, Basu, Amaresh, Modak, Mukund J
Format: Article
Language:English
Subjects:
Analytical, structural and metabolic biochemistry
Binding Sites
Biological and medical sciences
DNA Polymerase I - antagonists & inhibitors
DNA Polymerase I - metabolism
DNA Polymerase I - radiation effects
DNA, Bacterial - biosynthesis
Enzymes and enzyme inhibitors
Escherichia coli - metabolism
Fundamental and applied biological sciences. Psychology
Kinetics
Lysine
Peptide Fragments - isolation & purification
Pyridoxal Phosphate - pharmacology
Transferases
Trypsin
Ultraviolet Rays
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Summary:Inactivation of Escherichia coli DNA polymerase I by pyridoxal 5'-phosphate treatment results from its reactivity at multiple lysine residues. One of these residues, lysine-758, has been shown to be located at the substrate binding site in DNA polymerase I [Basu, A., & Modak, M. J. (1987) Biochemistry 26, 1704-1709]. We now demonstrate that lysine-635 is another important target of pyridoxylation; modification of this site results in decreased rates of DNA synthesis. Addition of template-primer with or without substrate deoxynucleoside triphosphate protects lysine-635 from pyridoxylation. Analysis of the initiation versus elongation phase of DNA synthesis by lysine-635-modified enzyme revealed that elongation of the DNA chain is severely affected by the lysine-635 modification. We therefore conclude that this lysine residue plays an important role in the processive mode of DNA synthesis by E. coli DNA polymerase I.
ISSN:0006-2960
1520-4995
DOI:10.1021/bi00418a011