Transition states of native and drug-resistant HIV-1 protease are the same

Kipp, D. Randal; Hirschi, Jennifer S.; Wakata, Aya; Goldstein, Harris; Schramm, Vern L.

doi:10.1073/pnas.1202808109

Cited by 25 publications

(44 citation statements)

References 54 publications

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“…As shown in Table 1, and in agreement with the conclusions derived from the analysis of the energetics of the alternative reaction paths, only the KIEs obtained from the GD mechanism are in qualitative and almost quantitative agreement with the experimental data. 33, 114 …”

Section: Resultsmentioning

confidence: 99%

“…As a consequence, a normal KIE is obtained (1.007), not in accordance with the experimental data of Schramm and co-workers. 33 …”

Section: Resultsmentioning

confidence: 99%

“…More recently, Schramm and co-workers proposed a model of transition state (TS) structure related with the decomposition of the GD intermediate, consistent with primary and secondary KIEs for native and I84V multidrug-resistant HIV-1 PR enzyme. 33 The conversion of the GD intermediate to the product complex should be rate limiting according to other kinetic studies. 17, 34, 35, 36 15 N KIEs were previously used by Meek and co-workers to elucidate the reaction mechanism of HIV-1 PR arriving to the conclusion that the protonation of the nitrogen atom of the amide leaving group should contribute to the rate limiting step.…”

Section: Introductionmentioning

confidence: 99%

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Dynamic and Electrostatic Effects on the Reaction Catalyzed by HIV-1 Protease

2016

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HIV-1 Protease (HIV-1 PR) is one of the three enzymes essential for the replication process of HIV-1 virus, which explains why it has been the main target for design of drugs against acquired immunodeficiency syndrome (AIDS). This work is focused on exploring the proteolysis reaction catalyzed by HIV-1 PR, with special attention to the dynamic and electrostatic effects governing its catalytic power. Free energy surfaces for all possible mechanisms have been computed in terms of potentials of mean force (PMFs) within hybrid QM/MM potentials, with the QM sub-set of atoms described at semiempirical (AM1) and DFT (M06-2X) level. The results suggest that the most favorable reaction mechanism involves formation of a gem-diol intermediate, whose decomposition into the product complex would correspond to the rate-limiting step. The agreement between the activation free energy of this step with experimental data, as well as kinetic isotope effects (KIEs), supports this prediction. The role of the protein dynamic was studied by protein isotope labelling in the framework of the Variational Transition State Theory. The predicted enzyme KIEs, also very close to the values measured experimentally, reveal a measurable but small dynamic effect. Our calculations show how the contribution of dynamic effects to the effective activation free energy appears to be below 1 kcal·mol−1. On the contrary, the electric field created by the protein in the active site of the enzyme emerges as being critical for the electronic reorganization required during the reaction. These electrostatic properties of the active site could be used as a mould for future drug design.

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Section: Resultsmentioning

confidence: 99%

“…As a consequence, a normal KIE is obtained (1.007), not in accordance with the experimental data of Schramm and co-workers. 33 …”

Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Dynamic and Electrostatic Effects on the Reaction Catalyzed by HIV-1 Protease

2016

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“…1). DHFR from Escherichia coli (EcDHFR) contains a number of mobile segments including the M20 (residues 9-24), FG (residues [116][117][118][119][120][121][122][123][124][125][126][127][128][129][130][131][132] and GH (residues 142-149) loops and switches between a closed and an occluded conformation during the catalytic cycle (Fig. 2).…”

Section: Introductionmentioning

confidence: 99%

Protein motions and dynamic effects in enzyme catalysis

Luk

Loveridge

Allemann

2015

Phys. Chem. Chem. Phys.

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The role of protein motions in promoting the chemical step of enzyme catalysed reactions remains a subject of considerable debate. Here, a unified view of the role of protein dynamics in dihydrofolate reductase catalysis is described. Recently the role of such motions has been investigated by characterising the biophysical properties of isotopically substituted enzymes through a combination of experimental and computational analyses. Together with previous work, these results suggest that dynamic coupling to the chemical coordinate is detrimental to catalysis and may have been selected against during DHFR evolution. The full catalytic power of Nature's catalysts appears to depend on finely tuning protein motions in each step of the catalytic cycle.

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“…18 Recently, the transition states of native and drug-resistant HIV-1 PR were shown to be the same. 19 Because both primary and secondary mutations are present in multidrug-resistant HIV-1 PR constructs that we previously analyzed by double electron-electron resonance (DEER) spectroscopy, 20, 21 where flap conformation and flexibility are altered, we hypothesize that these mutations individually and in combinations can modify HIV-1 PR flap conformational sampling.…”

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confidence: 99%

Elucidating a Relationship between Conformational Sampling and Drug Resistance in HIV-1 Protease

et al. 2013

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Enzyme targets in rapidly replicating systems, such as retroviruses, commonly respond to drug-selective pressure with mutations arising in the active site pocket that limit inhibitor effectiveness by introducing steric hindrance or by eliminating essential molecular interactions. However, these primary mutations are disposed to compromising pathogenic fitness. Emerging secondary mutations, which are often found outside of the binding cavity, may or can restore fitness while maintaining drug resistance. The accumulated drug-pressure selected mutations could have an indirect effect in the development of resistance, such as altering protein flexibility or the dynamics of protein-ligand interactions. Here, we show that accumulation of mutations in a drug-resistant variant, D30N/M36I/A71V, HIV-1 protease (HIV-1 PR) changes the fractional occupancy of the equilibrium conformational sampling ensemble. Correlations are made among populations of the conformational states; namely, closed-like, semi-open, and open-like, with inhibition constants, as well as kinetic parameters. Mutations that stabilize a closed-like conformation correlate with enzymes of lowered activity and with higher affinity for inhibitors, which is corroborated by a further increase in the fractional occupancy of the closed state upon addition of inhibitor or substrate-mimic. Cross resistance is found to correlate with combinations of mutations that increase the population of the open-like conformations at the expense of the closed-like state while retaining native-like occupancy of the semi-open population. These correlations suggest that at least three states are required in the conformational sampling model to establish the emergence of drug-resistance in HIV-1 PR. More importantly, these results shed light on a possible mechanism whereby mutations combine to impart drug resistance while maintaining catalytic activity.

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Transition states of native and drug-resistant HIV-1 protease are the same

Cited by 25 publications

References 54 publications

Dynamic and Electrostatic Effects on the Reaction Catalyzed by HIV-1 Protease

Dynamic and Electrostatic Effects on the Reaction Catalyzed by HIV-1 Protease

Protein motions and dynamic effects in enzyme catalysis

Elucidating a Relationship between Conformational Sampling and Drug Resistance in HIV-1 Protease

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