Inhibition of the RNase H activity of HIV reverse transcriptase by azidothymidylate

Cheng Keat Tan, Rigal Civil, A. Mohsin Mian, Antero G. So, Kathleen M. Downey

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Abstract

The effects of AZTMP and other nucleoside 5′-monophosphates on the RNA-dependent DNA polymerase and RNase H activities of a recombinant HIV reverse transcriptase have been investigated. Both activities are sensitive to inhibition by millimolar concentrations of AZTMP with MgCl2 as divalent cation activator. Substitution of Mn2+ for Mg2+ markedly potentiates the inhibition of RNase H activity by AZTMP, reducing the IC50 from 5 to 0.05 mM. In contrast, Mn2+ does not alter the sensitivity of the RNA-dependent DNA polymerase activity to inhibition by AZTMP. The inhibition of RNase H activity by AZTMP can be reversed by increasing concentrations of the substrate poly(A)/poly(dT), suggesting that AZTMP may compete with the substrate for binding at the active site of RNase H. Other nucleoside 5′-monophosphates do not inhibit RNase H in the presence of Mg2+. However, in the presence of Mn2+, deoxy- and dideoxynucleoside 5′-monophosphates that are complementary to the DNA strand of the heteroduplex substrate are somewhat inhibitory. The RNA-dependent DNA polymerase activity is a slightly inhibited by AZTMP and ddTMP in either Mg2+ or Mn2+, and substitution of Mn2+ for Mg2+ results in inhibition by ddAMP as well. Naturally occurring ribo- or deoxyribonucleoside 5′-monophosphates are not inhibitory at concentrations up to 5 mM. Since AZTTP inhibits the RNA-dependent DNA polymerase activity of HIV reverse transcriptase at nanomolar concentrations, it is unlikely that the inhibition of this activity by AZTMP plays a significant role in the antiviral effect of AZT. However, the inhibition of the RNase H activity by AZTMP, which can reach millimolar concentrations in vivo, may account for part of the sensitivity of the virus to AZT.

Original languageEnglish
Pages (from-to)4831-4835
Number of pages5
JournalBiochemistry
Volume30
Issue number20
StatePublished - Dec 1 1991
Externally publishedYes

ASJC Scopus subject areas

  • Biochemistry

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    Tan, C. K., Civil, R., Mohsin Mian, A., So, A. G., & Downey, K. M. (1991). Inhibition of the RNase H activity of HIV reverse transcriptase by azidothymidylate. Biochemistry, 30(20), 4831-4835.