Kinetics of Inhibition of Deoxynucleotide‐Polymerizing Enzyme Activities from Normal and Leukemic Human Cells by 9‐β‐D‐Arabinofuranosyladenine 5′‐Triphosphate and 1‐β‐D‐Arabinofuranosylcytosine 5′‐Triphosphate

Abstract

Both 9‐β‐D‐arabinofuranosyladenine 5′‐triphosphate (aATP) and 1‐β‐D‐arabinofuranosylcytosine 5′‐triphosphate (aCTP) inhibit terminal deoxynucleotidyl transferase and DNA polymerases α and β but not DNA polymerase γ from human cells. Inhibition of terminal deoxynucleotidyl transferase by both compounds is competitive with respect to either dGTP, dCTP, dATP, or dTTP and inhibition of DNA polymerases α and β is competitive with respect to dATP (for aATP) and dCTP (for aCTP). The inhibition constants for aATP for each enzyme are lower than the inhibition constants for aCTP implying that aATP has a stronger affinity for each enzyme than aCTP. In addition, the inhibition constants for aATP for each enzyme are about equal, suggesting that the drug binds to each enzyme with about the same affinity. The same is true for aCTP. Both compounds are non‐competitive inhibitors of terminal deoxynucleotidyl transferase with respect to initiator [(dA) 12–18 ], uncompetitive inhibitors of DNA polymerases α and β with respect to template/primer (activated DNA), and non‐competitive inhibitors of DNA polymerases α and β with respect to a non‐analogous substrate (dTTP). Inhibition of all three enzymes was reversed by simple dilution but not by additional enzyme, template/primer, initiator, or divalent cation. These results suggest that aATP and aCTP inhibit terminal deoxynucleotidyl transferase by competing with any substrate (dGTP, dCTP, dATP, or dTTP) for binding to the enzyme and DNA polymerases α and β by specifically competing with their analogs (dATP or dCTP) for binding to these enzymes.