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HIV-1 resistance to zidovudine: interactions between reverse transcriptase and RNA template.




 

Natl Conf Hum Retroviruses Relat Infect (1st). 1993 Dec 12-16;:136.

Polymerase activity of reverse transcriptase (RT) was studied under conditions limited to a single association of enzyme with a primertemplate (processive conditions). Polymerization under such conditions was compared to allowing multiple cycles of RT-primer- template (PT) association (non-processive conditions). These studies were undertaken because a change in zidovudine-triphosphate (ZDV-TP) binding has not been evident in RTs with mutations in codons 67, 70, 215, and 219. Recombinant wild type (HxB2) and mutant (with ZDV resistance mutations in codons 67, 70, 215, and 219) RT expression vectors were constructed. RTs were purified by L. Kohlstaedt and G. Lindwall in T. Steitz's lab. Extension of a 32P end-labelled DNA primer annealed to an RNA template was assessed by electrophoresis. Following preincubation of RT with 10- 20 fold excess of PT, DNA polymerization was initiated by the addition of magnesium (10 mM) and the 4 dNTPs (150 micromolar each). Heparin (2 mU/microliter) was added as a trap for dissociated RT (processive conditions). 3H-TTP incorporation into poly rA-oligo dT controlled for equivalent specific activities of the two different RTs. A greater accumulation of longer DNA products by the mutant than wild type RT was seen under processive conditions, both in the absence (7 expts.) And the presence (3 expts.) of ZDV-TP (10-25 micromolar). Syntheses by wild type and mutant RTs were identical under non-processive conditions. Further studies of these and other enzyme-template interactions, which may help elucidate the mechanism of ZDV resistance and suggest new approaches to RT inhibition, are underway.

Drug Resistance, Microbial Human HIV Infections/*DRUG THERAPY/VIROLOGY HIV-1/*DRUG EFFECTS Interferon Alfa-2b/ADMINISTRATION & DOSAGE/*THERAPEUTIC USE Zidovudine/ADMINISTRATION & DOSAGE/*THERAPEUTIC USE ABSTRACT



 




Information in this article was accurate in December 30, 1995. The state of the art may have changed since the publication date. This material is designed to support, not replace, the relationship that exists between you and your doctor. Always discuss treatment options with a doctor who specializes in treating HIV.