3rd International Workshop on HIV Drug Resistance 2-5 August 1994, Kauai, Hawaii, USA

MECHANISMS OF RESISTANCE OF HIV-1 RT TO NUCLEOSIDE AND NONNUCLEOSIDE INHIBITORS

Int Wkshop HIV Drug Res 1994 Aug 2-5;3:31 (abstract no. 30)

P.L. Boyer¹, C. Tantillo², J. Ding², A.L. Ferris¹, P. Clark³, E. Arnold² and S.H. Hughes^1
1ABL-Basic Research Program; ²Center for Advanced Biotechnology and Medicine and Rutgers University, Piscataway, NJ, USA; ³PRI/DynCorp, NCI-Frederick Cancer Research and Development Center, Frederick, MD, USA

There are two broad classes of inhibitors of HIV-1 RT: nucleosides and nonnucleosides. Treatment with either class of inhibitors results in drug resistance. We have developed E.coli expression plasmids that encode separate p51 and p66 subunits and have used these plasmids to produce HIV-1 RT with specific amino acid substitutions in one of the two subunits. Most of the mutations that confer resistance to nonnucleoside inhibitors cause resistance when present in the p66 subunit of the p66/p51 heterodimer; heterodimers that contained a wild-type p66 subunit and a mutant p51 subunit remained sensitive to the inhibitors. However, there was one mutation, E138K, that conferred drug resistance if, and only if, the mutation was present in the p51 subunit. Analysis of the three-dimensional structure of HIV-1 RT indicated that while the majority of the nonnucleoside drug binding pocket is formed by the p66 subunit, a small portion, including residue 138, is contributed by the p51 subunit.

We have studied two mutations, Leu 74Val and Glu89Gly, which confer resistance only to dideoxy nucleoside triphosphates. Neither of these amino acid positions are near the polymerase active site; however, both are in positions where they appear to interact with nucleic acid. Biochemical analyses suggest that both mutations affect the precise positioning of the template/prime and, as a consequence, affect the ability of the enzyme to discriminate between a deoxy and a dideoxy nucleoside triphosphate.

Research sponsored in part by the National Cancer Institute, DHHS, under contract No. NOI-CO-74101 with ABL and by the National Institute of General Medical Sciences.

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1994-08-02
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