Recent Advances in Understanding Biological GTP Hydrolysis through Molecular Simulation.

GTP hydrolysis is central to biology, being involved in regulating a wide range of cellular processes. However, the mechanisms by which GTPases hydrolyze this critical reaction remain controversial, with multiple mechanistic possibilities having been proposed based on analysis of experimental and computational data. In this mini-review, we discuss advances in our understanding of biological GTP hydrolysis based on recent computational studies and argue in favor of solvent-assisted hydrolysis as a conserved mechanism among GTPases. A concrete understanding of the fundamental mechanisms by which these enzymes facilitate GTP hydrolysis will have significant impact both for drug discovery efforts and for unraveling the role of oncogenic mutations.

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

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.

The role of ligand-gated conformational changes in enzyme catalysis.

Structural and biochemical studies on diverse enzymes have highlighted the importance of ligand-gated conformational changes in enzyme catalysis, where the intrinsic binding energy of the common phosphoryl group of their substrates is used to drive energetically unfavorable conformational changes in catalytic loops, from inactive open to catalytically competent closed conformations. However, computational studies have historically been unable to capture the activating role of these conformational changes. Here, we discuss recent experimental and computational studies, which can remarkably pinpoint the role of ligand-gated conformational changes in enzyme catalysis, even when not modeling the loop dynamics explicitly. Finally, through our joint analyses of these data, we demonstrate how the synergy between theory and experiment is crucial for furthering our understanding of enzyme catalysis.