The K65R Mutation in Human Immunodeficiency Virus Type 1 Reverse Transcriptase Exhibits Bidirectional Phenotypic Antagonism with Thymidine Analog Mutations

Parikh, Urvi M.; Bacheler, Lee T.; Koontz, Dianna; Mellors, John W.

doi:10.1128/jvi.80.10.4971-4977.2006

Cited by 116 publications

(87 citation statements)

References 46 publications

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“…Biochemical studies show that TAMs confer resistance to AZT and other NRTI by accelerating the rate at which the terminal AZT-MP (or other NRTI-MP) is removed through phosphorolytic cleavage (3,21). Both the K65R (this study; 25,29,35) and K70E (this study) mutant enzymes exhibit rates of ATP-or PPi-mediated phosphorolytic excision that are either significantly reduced or similar (in the case or PPi) to the WT enzyme. These data rule out the possibility that excision contributes to the NRTI resistance conferred by either of these mutations.…”

Section: Discussionmentioning

confidence: 99%

“…In this study, we provide the first insights into the molecular mechanism by which this mutation confers resistance to NRTI. Furthermore, we provide a direct comparison of the K70E mutation with K65R, a mutation that has also become more prevalent following the widespread use of TNV and ABC (17,25,32,37).…”

Section: Discussionmentioning

confidence: 99%

“…In this regard, previously uncommon mutations have become more prevalent among patients experiencing treatment failure. For example, since the introduction of NRTI, such as tenofovir (TNV) and abacavir (ABC), which select for the K65R mutation in HIV-1 RT, the incidence of this mutation has steadily increased in clinical databases (17,25,31,36).…”

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confidence: 99%

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Molecular Mechanism by Which the K70E Mutation in Human Immunodeficiency Virus Type 1 Reverse Transcriptase Confers Resistance to Nucleoside Reverse Transcriptase Inhibitors

Sluis‐Cremer

Sheen

Zelina

et al. 2007

Antimicrob Agents Chemother

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The K70E mutation in human immunodeficiency virus type 1 (HIV-1) reverse transcriptase (RT) has become more prevalent in clinical samples, particularly in isolates derived from patients for whom triple-nucleoside regimens that include tenofovir (TNV), abacavir, and lamivudine (3TC) failed. To elucidate the molecular mechanism by which this mutation confers resistance to these nucleoside RT inhibitors (NRTI), we conducted detailed biochemical analyses comparing wild-type (WT), K70E, and K65R HIV-1 RT. Pre-steady-state kinetic experiments demonstrate that the K70E mutation in HIV-1 RT allows the enzyme to discriminate between the natural deoxynucleoside triphosphate substrate and the NRTI triphosphate (NRTI-TP). Compared to the WT enzyme, K70E RT showed 2.1-, 2.3-, and 3.5-foldhigher levels of resistance toward TNV-diphosphate, carbovir-TP, and 3TC-TP, respectively. By comparison, K65R RT demonstrated 12.4-, 12.0-, and 13.1-fold-higher levels of resistance, respectively, toward the same analogs. NRTI-TP discrimination by the K70E (and K65R) mutation was primarily due to decreased rates of NRTI-TP incorporation and not to changes in analog binding affinity. The K65R and K70E mutations also profoundly impaired the ability of RT to excise 3 -azido-2 ,3 -dideoxythymidine monophosphate (AZT-MP) and other NRTI-MP from the 3 end of a chain-terminated primer. When introduced into an enzyme with the thymidine analog mutations (TAMs) M41L, L210W, and T215Y, the K70E mutation inhibited ATP-mediated excision of AZT-MP. Taken together, these findings indicate that the K70E mutation, like the K65R mutation, reduces susceptibility to NRTI by selectively decreasing NRTI-TP incorporation and is antagonistic to TAM-mediated nucleotide excision.

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

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Molecular Mechanism by Which the K70E Mutation in Human Immunodeficiency Virus Type 1 Reverse Transcriptase Confers Resistance to Nucleoside Reverse Transcriptase Inhibitors

Sluis‐Cremer

Sheen

Zelina

et al. 2007

Antimicrob Agents Chemother

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“…K65R is also a suppressor of AZT resistance by TAMs (6,36,43,59), and AZTMP excision activity is reduced in RTs containing this mutation (21,36,59). In contrast to the results obtained with M184V, the addition of K65R to the RT D67N/K70R/T215Y/K219Q background reduced AZTMP excision activity to similar extents on both DNA (Fig.…”

Section: Figmentioning

confidence: 99%

A Role of Template Cleavage in Reduced Excision of Chain-Terminating Nucleotides by Human Immunodeficiency Virus Type 1 Reverse Transcriptase Containing the M184V Mutation

Acosta-Hoyos

Matsuura

Meyer

et al. 2012

J Virol

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Resistance to nucleoside reverse transcriptase (RT) inhibitors is conferred on human immunodeficiency virus type 1 through thymidine analogue resistance mutations (TAMs) that increase the ability of RT to excise chain-terminating nucleotides after they have been incorporated. The RT mutation M184V is a potent suppressor of TAMs. In RT containing TAMs, the addition of M184V suppressed the excision of 3′-deoxy-3′-azidothymidine monophosphate (AZTMP) to a greater extent on an RNA template than on a DNA template with the same sequence. The catalytically inactive RNase H mutation E478Q abolished this difference. The reduction in excision activity was similar with either ATP or pyrophosphate as the acceptor substrate. Decreased excision of AZTMP was associated with increased cleavage of the RNA template at position −7 relative to the primer terminus, which led to increased primer-template dissociation. Whether M184V was present or not, RT did not initially bind at the −7 cleavage site. Cleavage at the initial site was followed by RT dissociation and rebinding at the −7 cleavage site, and the dissociation and rebinding were enhanced when the M184V mutation was present. In contrast to the effect of M184V, the K65R mutation suppressed the excision activity of RT to the same extent on either an RNA or a DNA template and did not alter the RNase H cleavage pattern. Based on these results, we propose that enhanced RNase H cleavage near the primer terminus plays a role in M184V suppression of AZT resistance, while K65R suppression occurs through a different mechanism.

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“…Also, clinical isolates and site-directed mutants harbouring TAMs were shown to display low-level decreased susceptibility to apricitabine in the presence or absence of the M184V mutation (Gu et al, 2006), suggesting that the effect of TAMs on apricitabine-mediated viral inhibition by apricitabine is not significant. Interestingly, both K65R and TAMs have been shown to exhibit bi-directional phenotypic and genotypic antagonism, which may explain the negative association of these mutations in genotype databases and the infrequent emergence of K65R in patients receiving zidovudine (Boucher et al, 2006;Parikh et al, 2006a;Parikh et al, 2006b).…”

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confidence: 99%

Kinetics of Inhibition of HIV Type 1 Reverse Transcriptase-Bearing NRTI-associated Mutations by Apricitabine Triphosphate

Frankel

Coutsinos

et al. 2007

Antivir Chem Chemother

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We wished to investigate the effects of various mutations in HIV-1 reverse transcriptase (RT) on biochemical inhibition by the active form of a novel nucleoside termed apricitabine. Accordingly, we studied the efficiency of chain-termination mediated by apricitabine triphosphate (TP) in cellfree assays that used either recombinant wild-type or mutated RTs. We also performed steady-statekinetics and primer-unblocking assays. Subtype C RTs were also analysed. The results showed that the K65R mutation in RT caused reductions in the efficiency of chain-termination of apricitabine-TP by increasing its K i . However, K65R did not affect rates of primer unblocking for apricitabine-TP. No significant differences were found between subtype C and subtype B RTs with regard to any of the parameters studied. Other mutations such as M184V, L74V and K103N had no effect on the efficiency of chain termination by apricitabine-TP. Thus, the mechanism of reduced susceptibility to apricitabine of viruses containing K65R in RT seems to be mediated exclusively through a reduction in binding or incorporation of apricitabine-TP. Unlike some other nucleoside analogues, increased excision of incorporated apricitabine does not seem to be a cause of resistance to apricitabine.

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The K65R Mutation in Human Immunodeficiency Virus Type 1 Reverse Transcriptase Exhibits Bidirectional Phenotypic Antagonism with Thymidine Analog Mutations

Cited by 116 publications

References 46 publications

Molecular Mechanism by Which the K70E Mutation in Human Immunodeficiency Virus Type 1 Reverse Transcriptase Confers Resistance to Nucleoside Reverse Transcriptase Inhibitors

Molecular Mechanism by Which the K70E Mutation in Human Immunodeficiency Virus Type 1 Reverse Transcriptase Confers Resistance to Nucleoside Reverse Transcriptase Inhibitors

A Role of Template Cleavage in Reduced Excision of Chain-Terminating Nucleotides by Human Immunodeficiency Virus Type 1 Reverse Transcriptase Containing the M184V Mutation

Kinetics of Inhibition of HIV Type 1 Reverse Transcriptase-Bearing NRTI-associated Mutations by Apricitabine Triphosphate

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