Effect of template secondary structure on the inhibition of HIV-1 reverse transcriptase by a pyridinone non-nucleoside inhibitor.
The importance of RNA secondary structure on HIV-1 reverse transcriptase catalyzed polymerization and on the potency of the pyridin-2-one inhibitor 3-(4,7-dichlorobenzoxazol-2-ylmethylamino)-5-ethyl-6-meth ylpyridin-2(1H)-one, L-697,661, were investigated by employing heteromeric primer-template systems. Our data revealed that a stem-loop hairpin secondary structure in the RNA template could lead to strong hindrance of reverse transcription in the reaction catalyzed by HIV-1 reverse transcriptase resulting in the build up of intermediate-length (pause) polymerization products. The presence of L-697,661 greatly enhanced the accumulation of the pause products suggesting that the rate of enzyme translocation from the pause product might be more potently inhibited than polymerization up to the pause site. Model experiments using a synthetic RNA template containing a stem-loop hairpin revealed that the inhibitory potency of L-697, 661 increased 2-fold upon polymerization to within four bases of the secondary structure. Inhibitor potency was enhanced over 6-fold when primer-extension proceeded through the duplex region of the stem-loop.
Olsen
D B
DB
Department of Biological Chemistry, Merck Research Laboratories, Merck & Co., West Point, PA 19486.
Carroll
S S
SS
Culberson
J C
JC
Shafer
J A
JA
Kuo
L C
LC
D000818:Animals; D000998:Antiviral Agents / Q000494:pharmacology*; D001483:Base Sequence; D001583:Benzoxazoles / Q000494:pharmacology*; D017931:DNA Primers; D004591:Electrophoresis, Polyacrylamide Gel; D005914:Globins / Q000235:genetics; D054303:HIV Reverse Transcriptase; D008969:Molecular Sequence Data; D015394:Molecular Structure; D009690:Nucleic Acid Conformation; D011728:Pyridones / Q000494:pharmacology*; D012367:RNA, Viral / Q000737:chemistry / Q000187:drug effects*; D011817:Rabbits; D018894:Reverse Transcriptase Inhibitors; D013698:Templates, Genetic
D000998:Antiviral Agents; D001583:Benzoxazoles; D017931:DNA Primers; D011728:Pyridones; D012367:RNA, Viral; D018894:Reverse Transcriptase Inhibitors; C076516:L-697661; D005914:Globins; D054303:HIV Reverse Transcriptase