Theoretical Modeling of Enzyme Reactions:  The Thermodynamics of Formation of Compound 0 in Horseradish Peroxidase

Abstract

In this paper, by using the perturbed matrix method (PMM) in combination with basic statistical mechanical relations both based on nanosecond time-scale molecular dynamics (MD) simulations, we quantitatively address the thermodynamics of compound 0 (Cpd 0) formation in horseradish peroxidase (HRP) enzyme. Our results, in the same trend of low-temperature experimental data, obtained in cryoenzymology studies indicate that such a reaction can be described essentially as a stepwise spontaneous process: a first step mechanically constrained, strongly exothermic proton transfer from the heme-H2O2 complex to the conserved His42, followed by a solvent-protein relaxation involving a large entropy increase. Critical evaluation of PMM/MD data also reveals the crucial role played by specific residues in the reaction pocket and, more in general, by the conformational fluctuations of the overall environment in physiological conditions.

The role of Arginine 38 in horseradish peroxidase enzyme revisited: A computational investigation

“…Similar conclusion was drawn in our previous paper (15), where it was found that after an exothermic and mechanically hindered proton transfer (from H 2 O 2 to His42), the reaction pocket at room temperature is in equilibrium between two conformations, differing essentially for the His42 position, and Cpd0 formation is therefore characterized by a large entropic contribution.…”

“…HRP structure (see Figure 2) is dominated by alphahelices with only two short anti-parallel beta-strands. This important feature, as already outlined in previous studies, (15,38) To this end, we report in Figure 4 the atom composition of the first two ED eigenvectors. In order to asses the actual differences between the internal fluctuations of the two systems we also calculated the average difference, and the related maximum error, between WT-HRP and R38L…”

“…The above residues are supposed to play a relevant role (3). As a matter of fact, Cpd0, a ferric-hydroperoxide anion, has been shown both experimentally (4)(5)(6)(7)(8)(9) and theoretically (10)(11)(12)(13)(14)(15) to be formed via deprotonation of H 2 O 2 by His42 acting as a base.…”

A Single-Site Mutation (F429H) Converts the Enzyme CYP 2B4 into a Heme Oxygenase: A QM/MM Study

“…3b,5e–f Details are given in the Supporting Information (SI), and those more specific to the paper are given here briefly. Since Cpd 0 was experimentally shown to exist in both the WT and MT enzymes, we started from the WT PDB 1SUO, 6a replaced the inhibitor by the OOH group, added missing hydrogen atoms, took care of the protonation states of acidic and basic residues, 3b,5d,f and followed with geometry optimization and molecular dynamic (MD) simulations. The F429H mutant was initially generated by replacing Phe429 by His, followed by minimization of the protein.…”

“…The so resulting WT and MT enzymes were both found to generate Cpd I with minor quantitative differences in the barriers of heterolytic cleavage (SI, Table S1). We therefore extended the MD simulation to 60 ns and extracted from the MD trajectories the most highly sampled conformational basins, 6b termed hereafter as, LWi and LMi, where L stands for long MD, W for the WT enzyme and M for the MT, i is the number of the basin. The WT enzyme prefers conformation LW1 and LW3.…”