HIV-1 reverse transcriptase plus-strand initiation exhibits preferential sensitivity to non-nucleoside reverse transcriptase inhibitors in vitro.
Non-nucleoside reverse transcriptase inhibitors (NNRTIs) are highly specific and potent allosteric inhibitors of human immunodeficiency virus type 1 (HIV-1) reverse transcriptase. NNRTIs inhibit reverse transcription in a substrate length-dependent manner in biochemical assays and in cell-based HIV-1 replication assays, suggesting a stochastic inhibitory mechanism. Surprisingly, we observed that NNRTIs potently inhibited plus-strand initiation in vitro under conditions in which little or no inhibition of minus-strand DNA synthesis was observed. In assays that recapitulated the initiation of plus-strand DNA synthesis, greater inhibition was observed with an RNA PPT primer than with a DNA primer of corresponding sequence and with wild-type reverse transcriptase but not with NNRTI-resistant enzymes. Structural elements that dictate sensitivity to NNRTIs were revealed using modified plus-strand initiation substrates. The data presented here suggest that specific inhibition of plus-strand initiation may be an important mechanism by which NNRTIs block HIV-1 replication.
Grobler
Jay A
JA
Department of Antiviral Research, Merck Research Laboratories, West Point, Pennsylvania 19486, USA. jay_grobler@merck.com
Dornadula
Geetha
G
Rice
Michele R
MR
Simcoe
Amy L
AL
Hazuda
Daria J
DJ
Miller
Michael D
MD
D000480:Alkynes; D019380:Anti-HIV Agents / Q000494:pharmacology; D048588:Benzoxazines; D001583:Benzoxazoles / Q000494:pharmacology; D003521:Cyclopropanes; D004247:DNA / Q000737:chemistry; D004305:Dose-Response Relationship, Drug; D024882:Drug Resistance, Viral; D004791:Enzyme Inhibitors / Q000494:pharmacology; D054303:HIV Reverse Transcriptase / Q000378:metabolism*; D066298:In Vitro Techniques; D020128:Inhibitory Concentration 50; D008024:Ligands; D008957:Models, Genetic; D010078:Oxazines / Q000494:pharmacology; D011728:Pyridones / Q000494:pharmacology; D018894:Reverse Transcriptase Inhibitors / Q000494:pharmacology*; D013269:Stochastic Processes; D014158:Transcription, Genetic; D014779:Virus Replication
D000480:Alkynes; D019380:Anti-HIV Agents; D048588:Benzoxazines; D001583:Benzoxazoles; D003521:Cyclopropanes; D004791:Enzyme Inhibitors; D008024:Ligands; D010078:Oxazines; D011728:Pyridones; D018894:Reverse Transcriptase Inhibitors; C076516:L-697661; D004247:DNA; D054303:HIV Reverse Transcriptase; C098320:efavirenz