Steady state kinetics and inhibition of HIV-1 reverse transcriptase by a non-nucleoside dipyridodiazepinone, BI-RG-587, using a heteropolymeric template

Nucleic Acids Res. 1991 Jun 11;19(11):3035-9. doi: 10.1093/nar/19.11.3035.

Abstract

Steady state kinetics and inhibition by a dipyridodiazepinone of the reverse transcriptase from human immunodeficiency virus type 1 (HIV) were studied using a heteropolymeric RNA template with a sequence from the authentic initiation site on the HIV genome. For addition of the first deoxynucleotide to primer, kcat/KM is 0.05 (nM-min)-1 and KM is 10 nM. When all 4 deoxynucleotide triphosphates are present and processive synthesis occurs, catalysis is less efficient; kcat/KM = .0077 (nM-min)-1 and KM = 100 nM for dATP. These results are consistent with a rate determining conformation change involved in translocation of the enzyme along the template. Inhibition by the dipyridodiazepinone BI-RG-587 is noncompetitive with respect to both nucleotide and template-primer; this compound decreases Vmax but does not affect KM. Thus, this inhibitor binds to a site distinct from the substrate binding sites with Ki of 220 nM. Inhibition by BI-RG-587 results in a uniform decrease in amount of products of all lengths rather than a shift from longer to shorter products, suggesting the inhibitor does not affect processivity of reverse transcriptase.

MeSH terms

  • Base Sequence
  • Electrophoresis, Polyacrylamide Gel
  • HIV-1 / enzymology*
  • Kinetics
  • Molecular Sequence Data
  • Nevirapine
  • Pyridines / pharmacology*
  • RNA, Viral / genetics
  • Reverse Transcriptase Inhibitors*
  • Templates, Genetic

Substances

  • Pyridines
  • RNA, Viral
  • Reverse Transcriptase Inhibitors
  • Nevirapine