Structural and Thermodynamic Basis of Amprenavir/Darunavir and Atazanavir Resistance in HIV-1 Protease with Mutations at Residue 50

Mittal, Sakshi; Bandaranayake, Rajinthna M.; King, Nancy M. P.; Prabu-Jeyabalan, M.; Nalam, M.N.L.; Nalivaika, E.A.; Yilmaz, Neşe Kurt; Schiffer, Celia A.

doi:10.1128/jvi.03486-12

Cited by 29 publications

(35 citation statements)

References 36 publications

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“…Previously, we showed that the I50V/A71V protease has decreased vdW interactions with the protease inhibitors APV and DRV compared with WT (0.61 and 1.98 kcal/mol, respectively), mainly due to the loss of a methyl group interacting with the sulfonyl moiety in APV/DRV (39). The coevolved I50V/A71V LP1′F and I50V/A71V PP5′L structures have more vdW contacts, and I50V/A71V RP4′S has more hydrogen bonds compared with WT WT complex.…”

Section: Discussionmentioning

confidence: 97%

Structural basis and distal effects of Gag substrate coevolution in drug resistance to HIV-1 protease

Özen

Lin

Yilmaz

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

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Drug resistance mutations in response to HIV-1 protease inhibitors are selected not only in the drug target but elsewhere in the viral genome, especially at the protease cleavage sites in the precursor protein Gag. To understand the molecular basis of this proteasesubstrate coevolution, we solved the crystal structures of drug resistant I50V/A71V HIV-1 protease with p1-p6 substrates bearing coevolved mutations. Analyses of the protease-substrate interactions reveal that compensatory coevolved mutations in the substrate do not restore interactions lost due to protease mutations, but instead establish other interactions that are not restricted to the site of mutation. Mutation of a substrate residue has distal effects on other residues' interactions as well, including through the induction of a conformational change in the protease. Additionally, molecular dynamics simulations suggest that restoration of active site dynamics is an additional constraint in the selection of coevolved mutations. Hence, protease-substrate coevolution permits mutational, structural, and dynamic changes via molecular mechanisms that involve distal effects contributing to drug resistance.HIV-1 protease | drug resistance | coevolution | crystallography | active site dynamics

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Section: Discussionmentioning

confidence: 97%

Structural basis and distal effects of Gag substrate coevolution in drug resistance to HIV-1 protease

Özen

Lin

Yilmaz

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

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“…44 This was not the case for TPV, where a reduction in ΔΔH was reported for the same mutation. 44 Mutation A71V, which tends to appear together with I50V, 12 is known to compensate for the loss of viral fitness due to primary drug resistance-associated mutation. 83 An ITC study of I50V+A71V double mutant 12 suggested an increase in affinity toward ATV as a result of an increase in entropy which compensated the increase in enthalpy.…”

Section: Journal Of Chemical Theory and Computationmentioning

confidence: 99%

“…It also has been suggested to cause resistance toward Indinavir (IDV), 9,11 and at the same time, there is evidence pointing that this mutation has been associated with sensitivity toward Atazanavir (ATV) 12,13 and Tipranavir (TPV).…”

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confidence: 99%

Consistent Prediction of Mutation Effect on Drug Binding in HIV-1 Protease Using Alchemical Calculations

Bastys

Gapsys

Doncheva

et al. 2018

J. Chem. Theory Comput.

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Despite a large number of antiretroviral drugs targeting HIV-1 protease for inhibition, mutations in this protein during the course of patient treatment can render them inefficient. This emerging resistance inspired numerous computational studies of the HIV-1 protease aimed at predicting the effect of mutations on drug binding in terms of free binding energy ΔG, as well as in mechanistic terms. In this study, we analyze ten different protease-inhibitor complexes carrying major resistance-associated mutations (RAMs) G48V, I50V, and L90M using molecular dynamics simulations. We demonstrate that alchemical free energy calculations can consistently predict the effect of mutations on drug binding. By explicitly probing different protonation states of the catalytic aspartic dyad, we reveal the importance of the correct choice of protonation state for the accuracy of the result. We also provide insight into how different mutations affect drug binding in their specific ways, with the unifying theme of how all of them affect the crucial drug binding regions of the protease. ■ INTRODUCTIONHIV-1 (human immunodeficiency virus-1, further denoted as HIV) has caused a global epidemic that affects approximately 37 million people worldwide.1 There is no vaccine or cure available against HIV, but antiretroviral therapy (ART) is recommended for every infected individual 1 to suppress the virus. Success of ART has led to a near-normal life expectancy of HIV infected patients.2 Nevertheless, resistance toward drugs remains a major issue, making it necessary to switch therapy during the course of treatment of a single patient. 1One of the main targets of ART is the HIV protease, a protein responsible for cleaving HIV polyproteins during virus maturation ( Figure 1a). HIV protease is a homodimer with two 99 residues long subunits. The major structural components of the binding pocket of this protein are the active site at its bottom, which is followed in sequence by a loop and a short β-sheet (residues 26−32) that constitute the side of the pocket together with the so-called 80s loop (residues 79−84) ( Figure 1a). On the top the pocket is covered by the so-called flap region. In the framework of ART, HIV protease can be inhibited by protease inhibitors (PIs) (Figure 1b), a class of competitive inhibitors which are currently recommended as second-and third-line ART treatment.1 Specific substitutions in 13 different positions in the protease are considered to be major resistanceassociated mutations (RAMs). 3I50V is a RAM toward protease inhibitors Lopinavir (LPV) 4,5 and Darunavir (DRV) 6,7 and has been associated with particularly strong resistance to Amprenavir (APV) 8−10 and correspondingly to its prodrug Fosamprenavir (FPV). It also has been suggested to cause resistance toward Indinavir (IDV), 9,11 and at the same time, there is evidence pointing that this mutation has been associated with sensitivity toward Atazanavir (ATV) 12,13 and Tipranavir (TPV).14 I50 is located in the protease flap region (Figure 1a). This flap is...

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“…WT NL4-3 HIV-1 protease had a Michaelis-Menten constant, Km, of 71 ± 7 M for cleaving this substrate, which was similar to that of I84V and V82I variants (66 ± 4 and 62 ± 4 M, respectively). The primary resistance mutation I50V is known to reduce catalytic activity, 31 and while I50V is still catalytically active, the Km value is beyond the limit of detection for this assay. A71V, a compensatory mutation that is far from the active site and almost always observed with I50V, restores the functionality to WT level (Km = 73 ± 9 M), as previously reported.…”

Section: Enzymatic Activity Of Protease Variantsmentioning

confidence: 99%

“…Mutations at I50 are often selected together with A71V mutation, which is distal from the active site but compensates for the loss of enzymatic fitness 30 . Thermodynamics and structural studies of DRV binding to I50V/L and A71V mutations have revealed significant loss of van der Waals (vdW) contacts between the inhibitor and protease underlying loss of binding affinity 31 .…”

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confidence: 99%

Structural Adaptation of Darunavir Analogs Against Primary Resistance Mutations in HIV-1 Protease

Lockbaum

Leidner

Rusere

et al. 2018

Preprint

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HIV-1 protease is one of the prime targets of agents used in antiretroviral therapy against HIV. However, under selective pressure of protease inhibitors, primary mutations at the active site weaken inhibitor binding to confer resistance. Darunavir (DRV) is the most potent HIV-1 protease inhibitor in clinic; resistance is limited, as DRV fits well within the substrate envelope. Nevertheless, resistance is observed due to hydrophobic changes at residues including I50, V82 and I84 that line the S1/S1' pocket within the active site. Through enzyme inhibition assays and a series of 12 crystal structures, we interrogated susceptibility of DRV and two potent analogs to primary S1' mutations. The analogs had modifications at the hydrophobic P1' moiety to better occupy the unexploited space in the S1' pocket where the primary mutations were located. Considerable losses of potency were observed against protease variants with I84V and I50V mutations for all three inhibitors. The crystal structures revealed an unexpected conformational change in the flap region of I50V protease bound to the analog with the largest P1' moiety, indicating interdependency between the S1' subsite and the flap region. Collective analysis of protease-inhibitor van der Waals (vdW) interactions in the crystal structures using principle component analysis indicated I84V mutation underlying the largest variation in the vdW contacts. Interestingly, the principle components were able to distinguish inhibitor identity and relative potency solely based on vdW interactions of active site residues in the crystal structures. Our results reveal the interplay between inhibitor P1' moiety and primary S1' mutations, as well as suggesting a novel method for distinguishing the interdependence of resistance through principle component analyses.

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Structural and Thermodynamic Basis of Amprenavir/Darunavir and Atazanavir Resistance in HIV-1 Protease with Mutations at Residue 50

Cited by 29 publications

References 36 publications

Structural basis and distal effects of Gag substrate coevolution in drug resistance to HIV-1 protease

Structural basis and distal effects of Gag substrate coevolution in drug resistance to HIV-1 protease

Consistent Prediction of Mutation Effect on Drug Binding in HIV-1 Protease Using Alchemical Calculations

Structural Adaptation of Darunavir Analogs Against Primary Resistance Mutations in HIV-1 Protease

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