Different Enzyme Conformations Induce Different Mechanistic Traits in HIV‐1 Protease

Abstract

The influence of the dynamical flexibility of enzymes on reaction mechanisms is a cornerstone in biological sciences. In this study, we aim to 1) study the convergence of the activation free energy by using the first step of the reaction catalysed by HIV-1 protease as a case study, and 2) provide further evidence for a mechanistic divergence in this enzyme, as two different reaction pathways were seen to contribute to this step. We used quantum mechanics/molecular mechanics molecular dynamics simula-tions, on four different initial conformations that led to different barriers in a previous study. Despite the sampling, the four activation free energies still spanned a range of 5.0 kcal • mol À 1 . Furthermore, the new simulations did confirm the occurrence of an unusual mechanistic divergence, with two different mechanistic pathways displaying equivalent barriers. An active-site water molecule is proposed to influence the mechanistic pathway.

Rate-enhancing PETase mutations determined through DFT/MM molecular dynamics simulations

“…This observation agrees with several studies of the dependence between the activation free energy and the reactant structure. It has been shown in several different systems, such as the HIV-1 protease, 39,40 α-amylase, 41,42 2′-hydroxyphenyl-2-sulfinate desulfinase, 43 or the human transmembrane protease serine 2 44 that the activation barrier depends critically on the fine pre-organization of the active site. In an enoyl-thioester reductase, it was experimentally shown that a mutation causing a displacement of 1 Å in the substrate caused a change of six orders of magnitude in forming an otherwise undetectable side-product.…”

Unraveling the Reaction Mechanism of Russell’s Viper Venom Factor X Activator: A Paradigm for the Reactivity of Zinc Metalloproteinases?

“…In the case of aspartic proteases, the existence of a pair of active-site Asp residues, one neutral and one negative, allows overcoming the burden of deprotonating the very basic water molecule. While one of the Asp residues deprotonates the hydrolytic water molecule, the second Asp residue protonates the very basic aliphatic oxyanion that is generated by the water attack on the substrate’s carbonyl so the burden of the deprotonation is compensated by the simultaneous very exothermic protonation of the growing oxyanion. , In some cases, the protonation and deprotonation can be carried out by a single Asp residue …”

“…The latter represents an average over all proteins in the crystal and thus is the most important and informative of all structures. Our experience is that the x-ray structure can provide results similar to ensemble averages because it is an ensemble average itself. , …”

“…Our experience is that the x-ray structure can provide results similar to ensemble averages because it is an ensemble average itself. 76,77 QM/MM Model. The minimized structure of the proteinsubstrate complex was loaded on the molUP plugin 78 of the VMD program package, which provides a full-featured graphical user interface (GUI) to the software Gaussian.…”

“…99,102 In some cases, the protonation and deprotonation can be carried out by a single Asp residue. 103 The strategy of RVV-X for generating the nucleophile is to coordinate the water molecule to the Zn 2+ cofactor, lowering the water pK a through the much stronger coordination of the hydroxide ion in relation to the neutral water. The water deprotonation molecule moves the negative charge from the non-Zn 2+ -coordinated Glu140 to the Zn 2+ -coordinated water molecule, with an obvious electrostatic gain.…”