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. 2013 Mar;5(3):174-81.
doi: 10.1038/nchem.1559. Epub 2013 Jan 27.

Snapshot of the Equilibrium Dynamics of a Drug Bound to HIV-1 Reverse Transcriptase

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Free PMC article

Snapshot of the Equilibrium Dynamics of a Drug Bound to HIV-1 Reverse Transcriptase

Daniel G Kuroda et al. Nat Chem. .
Free PMC article

Abstract

The anti-AIDS drug rilpivirine undergoes conformational changes to bind HIV-1 reverse transcriptase (RT), which is an essential enzyme for the replication of HIV. These changes allow it to retain potency against mutations that otherwise would render the enzyme resistant. Here we report that water molecules play an essential role in this binding process. Femtosecond experiments and theory expose the molecular level dynamics of rilpivirine bound to HIV-1 RT. Two nitrile substituents, one on each arm of the drug, are used as vibrational probes of the structural dynamics within the binding pocket. Two-dimensional vibrational echo spectroscopy reveals that one nitrile group is unexpectedly hydrogen-bonded to a mobile water molecule, not identified in previous X-ray structures. Ultrafast nitrile-water dynamics are confirmed by simulations. A higher (1.51 Å) resolution X-ray structure also reveals a water-drug interaction network. Maintenance of a crucial anchoring hydrogen bond may help retain the potency of rilpivirine against pocket mutations despite the structural variations they cause.

Conflict of interest statement

The authors declare no competing financial interests.

Figures

Figure 1
Figure 1
Experimental linear IR spectra of rilpivirine in solution and in complex to the different RT enzymes. (a) FTIR and fits of inhibitor in methanol (black circles, green line), THF (black squares, red line), and DMSO (black triangles, blue line). (b) FTIR of the rilpivirine complex to the WT-RT in a single crystal (black squares), and its fit (redline, dash blue and dash green), (c) FTIR of the rilpivirine complex to the WT-RT in solution (black squares), (d) FTIR of the rilpivirine complex to the mutant M2-RT (Y181C/K103N) in solution (black squares), (e) FTIR of the rilpivirine complex to the mutant M1-RT (L100I/K103N) in solution (black squares), and their corresponding fits (red, dash blue, and dashed green lines).
Figure 2
Figure 2
Absorptive 2D IR spectra of the two investigated mutant-rilvipirine complexes at different waiting times. The left and right columns correspond to the spectra of M1-RT (L100I/K103N)/rilpivirine and M2-RT (Y181C/K103N)/rilpivirine, respectively. The waiting time of each spectrum is indicated in the figure.
Figure 3
Figure 3
Experimental peak shift decays extracted from 2D-IR data as a function of waiting time. Left and right columns correspond to the M1-RT (L100I/K103N)/rilpivirine and to the M2-RT (Y181C/K103N)/rilpivirine complexes, respectively. Panels (a) and (c) show the peak shift of the total signal and panels (b) and (d) to the peak shift with water signal subtracted. Red lines correspond to their fit as mentioned in the text.
Figure 4
Figure 4
Calculated radial distribution function g(r) highlighting differences between the two nitrile groups of rilpivirine. The radial distribution functions between the nitrogen of the nitrile group and the oxygen of water for: (a) benzonitrile arm and (b) cinnamonitrile arm of the inhibitor. Black, blue, and red lines correspond to the WT, M1-RT, and M2-RT enzymes, respectively.
Figure 5
Figure 5
Snapshot of rilpivirine in the NNRTI-binding pocket as observed in our MD simulations and X-ray crystal structure. Left and middle panels correspond to the different views of the pocket from the MD simulation where cyan, blue, red, and white correspond to carbon, nitrogen, oxygen, and hydrogen atoms, respectively, and the grey surface to the protein. Right panel corresponds to the thermal ellipsoid representation of rilpivirine in the NNRTI-binding pocket; the ellipsoids represent anisotropic thermal parameters of individual atoms that were refined using 1.51 Å resolution X-ray diffraction data. Two water molecules that interact with rilpivirine are represented as red ellipsoids.
Figure 6
Figure 6
Simulated absorptive 2D-IR spectra and peak shift dynamics of WT-RT/rilpivirine complex with (right column) and without (left column) Gaussian noise for different waiting times as indicated in the Figure. Top left panel peak shift dynamic calculated directly from the response functions. Top right panel peak shift dynamics calculated from a window in the simulated 2D IR spectrum. Squares and red line corresponds to simulations with and without noise, respectively. Parameters are presented in the Supporting Information (Table S5). No 2D-IR cross peak between the two nitrile transitions is located in the spectra for any of the mutant/drug complexes either in theory or experiments.

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