How the HIV-1 Reverse Transcriptase Mutations K65R, M184V and K65R + M184V Produce Resistance to NRTIs

There are two known mechanisms by which HIV-1 reverse transcriptase becomes resistant to nucleoside/nucleotide reverse transcriptase inhibitors (NRTIs): via reduced binding and incorporation, referred to as discrimination, and via increased NRTI excision after incorporation. We can measure discrimination using steady-state enzyme kinetics (Ki/Km), whereas excision after incorporation can be measured based on the depletion of adenosine triphosphate (ATP) or pyrophosphate and the generation of new deoxynucleotide triphosphates (dNTPs).

The reverse transcriptase resistance mutations K65R and M184V occur individually in HIV-1, as well as in combination. These mutations each increase phenotypic susceptibility to zidovudine (AZT, Retrovir), while the M184V mutation also increases susceptibility to tenofovir (TDF, Viread). HIV-1 carrying K65R has reduced susceptibility to most NRTIs, except zidovudine. M184V mutants have reduced susceptibility to lamivudine (3TC, Epivir) and emtricitabine (FTC, Emtriva) and may also contribute to abacavir (ABC, Ziagen) and didanosine (ddI, Videx) resistance. When M184V is added to K65R, susceptibility to tenofovir, zidovudine and stavudine (d4T, Zerit) increases slightly relative to K65R alone, whereas susceptibility to lamivudine, emtricitabine, didanosine and abacavir decreases.

In this in vitro enzymatic study, investigators from Gilead Sciences, Inc., demonstrated that the K65R, M184V and K65R + M184V mutations decreased the binding and incorporation of most NRTIs (indicated by higher Ki/Km values relative to wild type).1 However, reverse transcriptase with the M184V mutation alone showed no change in the rate of tenofovir, stavudine and zidovudine discrimination. The K65R + M184V combination resulted in additive resistance to didanosine and abacavir at the level of discrimination, but increased sensitivity relative to K65R alone for tenofovir, stavudine and zidovudine. Sensitization to zidovudine in the presence of K65R + M184V is likely caused by decreased excision.

Altered Binding or Incorporation of N(t)RTIs
Slide by J.K. Ly; reprinted with permission. Click here to download the complete slide presentation.

Reverse transcriptase harboring the K65R and K65R + M184V mutations showed decreased tenofovir and zidovudine excision, in addition to decreased binding and incorporation.

Altered Excision of N(t)RTIs (ATP-Mediated)
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Slide by J.K. Ly; reprinted with permission. Click here to download the complete slide presentation.

The authors concluded that for most NRTIs the K65R, M184V and K65R + M184V reverse transcriptase mutations primarily disrupt NRTI binding and incorporation. These findings correlate with the drug susceptibilities of these NRTIs in phenotypic assays. However, with zidovudine, increased discrimination by K65R and K65R + M184V is counteracted by decreased excision caused by these mutations, resulting in wild-type phenotypic susceptibility. As shown by these findings, it is important to consider both enzymatic reactions -- discrimination and excision -- when determining the mechanism of resistance caused by specific resistance mutations.


  1. Ly JK, Margot NA, Macarthur HL, Hung M, Miller MD, White KL. Resistance mechanisms of HIV-1 reverse transcriptase mutants K65R, M184V, and K65R+M184V to NRTIs. In: Program and abstracts of the 46th Interscience Conference on Antimicrobial Agents and Chemotherapy; September 27-30, 2006; San Francisco, Calif. Abstract H-1670.
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