Mechanism of selective inhibition of 3' to 5' exonuclease activity of Escherichia coli DNA polymerase I by nucleoside 5'-monophosphates

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Abstract
The 3'' to 5'' exonuclease activity of E. coli DNA polymerase I can be selectively inhibited by nucleoside 5''-monophosphates, but the DNA polymerase activity is not inhibited. The results of kinetic studies show that nucleotides containing a free 3''-hydroxy group and a 5''-phosphoryl group are competitive inhibitors of the 3'' to 5'' exonuclease. Previous studies by others demonstrated a binding site for nucleoside 5''-monophosphates on DNA polymerase I. The Kdissoc [dissociation constant] values for nucleoside 5''-monophosphates determined in that study are comparable to the Ki [inhibition constant] values determined in the present study, suggesting that the specific binding site for nucleoside 5''-monophosphates represents the inhibitor site of the 3'' to 5'' exonuclease activity. Thus, the binding site for nucleoside 5''-monophosphates on DNA polymerase I may represent the product site of the 3'' to 5'' exonuclease activity; the primer terminus site for the 3'' to 5'' exonuclease activity is distinct from the primer terminus site for the polymerase activity; and nucleoside 5''-monophosphates bind at the primer terminus site for the 3'' to 5'' exonuclease activity.