Conformational diversity induces nanosecond-timescale chemical disorder in the HIV-1 protease reaction pathway†

Abstract

One enzyme, one substrate, but two different reaction mechanisms: HIV-1 protease follows different reaction mechanisms depending on its instantaneous conformation.

Abstract

The role of conformational diversity in enzyme catalysis has been a matter of analysis in recent studies. Pre-organization of the active site has been pointed out as the major source for enzymes' catalytic power. Following this line of thought, it is becoming clear that specific, instantaneous, non-rare enzyme conformations that make the active site perfectly pre-organized for the reaction lead to the lowest activation barriers that mostly contribute to the macroscopically observed reaction rate. The present work is focused on exploring the relationship between structure and catalysis in HIV-1 protease (PR) with an adiabatic mapping method, starting from different initial structures, collected from a classical MD simulation. The first, rate-limiting step of the HIV-1 PR catalytic mechanism was studied with the ONIOM QM/MM methodology (B3LYP/6-31G(d):ff99SB), with activation and reaction energies calculated at the M06-2X/6-311++G(2d,2p):ff99SB level of theory, in 19 different enzyme:substrate conformations. The results showed that the instantaneous enzyme conformations have two independent consequences on the enzyme's chemistry: they influence the barrier height, something also observed in the past in other enzymes, and they also influence the specific reaction pathway, which is something unusual and unexpected, challenging the “one enzyme–one substrate–one reaction mechanism” paradigm. Two different reaction mechanisms, with similar reactant probabilities and barrier heights, lead to the same gem-diol intermediate. Subtle nanosecond-timescale rearrangements in the active site hydrogen bonding network were shown to determine which reaction the enzyme follows. We named this phenomenon chemical disorder. The results make us realize the unexpected mechanistic consequences of conformational diversity in enzymatic reactivity.

Notes

†Electronic supplementary information (ESI) available: Methodology used in the initial model and MM-MD simulations; Fig. S1 – energy distribution of the NPT ensemble generated by the MM-MD calculations and of the microstates extracted for the QM/MM calculations; Fig. S2 – correlations between six selected active site distances from reactant structures and the corresponding activation barriers; Fig. S3 – correlations between six selected active site distances from transition state structures and the corresponding activation barriers (PDF). Geometries of the optimized stationary points (ZIP file). See DOI: 10.1039/c9sc01464k