Inhibition of human immunodeficiency virus 1 reverse transcriptase by 3'-azidothymidine triphosphate.
In the presence of oligo(dT).poly(rA) as primer-template, 3'-azidothymidine triphosphate (N3'(3)-ddTTP) is a substrate for human immunodeficiency virus 1 reverse transcriptase with an apparent Km value of 3.0 microM. This compares with an apparent Km for thymidine monophosphate (dTMP) incorporation of 2.5 microM. The apparent Vmax value for 3'-azidothymidine monophosphate (N3'(3)-ddTMP) incorporation is 50 times lower than that of dTMP incorporation. Kinetic analysis of the inhibition of reverse transcriptase by N3'(3)-ddTTP shows competitive inhibition with thymidine triphosphate (dTTP) with a Ki of 41 nM and an uncompetitive pattern of inhibition with template-primer having a Ki of 140 nM. This indicates incorporation of the analogue into the primer and inhibition of the enzyme by formation of a dead-end complex. The 3'-azidothymidine-terminated primer-template [N3'(3)-ddT-(dT)15.poly(rA)] is a potent competitive inhibitor versus primer-template with a Ki of 2.4 nM and shows mixed-type inhibition against dTTP with a Ki of 8 nM. The low inhibition constant for this chain-terminated primer suggests that such oligonucleotides can act as potent inhibitors of enzyme catalysis.
Max-Planck-Institut für Experimentelle Medizin, Abteilung Chemie, Göttingen, Federal Republic of Germany.
O
O
Heidenreich
M
M
Kruhøffer
F
F
Grosse
F
F
Eckstein
D001665:Binding Sites / Q000187:drug effects; D002384:Catalysis / Q000187:drug effects; D015497:HIV-1 / Q000201:enzymology*; D007700:Kinetics; D012194:RNA-Directed DNA Polymerase / Q000737:chemistry; D018894:Reverse Transcriptase Inhibitors; D013379:Substrate Specificity; D013938:Thymidine Monophosphate / Q000737:chemistry / Q000494:pharmacology; D013942:Thymine Nucleotides / Q000737:chemistry / Q000494:pharmacology; D015215:Zidovudine / Q000737:chemistry / Q000494:pharmacology*
D018894:Reverse Transcriptase Inhibitors; D013942:Thymine Nucleotides; D013938:Thymidine Monophosphate; D015215:Zidovudine; D012194:RNA-Directed DNA Polymerase; C024157:thymidine 5'-triphosphate