Multiscale Simulations Give Insight into the Hydrogen In and Out Pathways of [NiFe]-Hydrogenases from Aquifex aeolicus and Desulfovibrio fructosovorans

Abstract

[NiFe]-hydrogenases catalyze the cleavage of molecular hydrogen into protons and electrons and represent promising tools for H2-based technologies such as biofuel cells. However, many aspects of these enzymes remain to be understood, in particular how the catalytic center can be protected from irreversible inactivation by O2. In this work, we combined homology modeling, all-atom molecular dynamics, and coarse-grain Brownian dynamics simulations to investigate and compare the dynamic and mechanical properties of two [NiFe]-hydrogenases: the soluble O2-sensitive enzyme from Desulfovibrio fructosovorans, and the O2-tolerant membrane-bound hydrogenase from Aquifex aeolicus. We investigated the diffusion pathways of H2 from the enzyme surface to the central [NiFe] active site, and the possible proton pathways that are used to evacuate hydrogen after the oxidation reaction. Our results highlight common features of the two enzymes, such as a Val/Leu/Arg triad of key residues that controls ligand migration and substrate access in the vicinity of the active site, or the key role played by a Glu residue for proton transfer after hydrogen oxidation. We show specificities of each hydrogenase regarding the enzymes internal tunnel network or the proton transport pathways.

Fold and flexibility: what can proteins' mechanical properties tell us about their folding nucleus?

“…As shown in previous mechanical studies done on members of the globin family [39] and homologous hydrogenases [40], proteins with similar folds also present similar rigidity profiles. For each protein fold that was studied, we define the consensus nucleus as the subset of conserved positions along the protein sequence that present rigidity peaks in all (or most of, in the case of largest alignments) the rigidity profiles of the structures that were studied.…”

“…In order to investigate them on a residue level, we developed the PROPHET (Probing Protein Heterogeneity) program [37], which combines a coarse-grain (CG) elastic network representation and Brownian dynamics (BD). Studies using PROPHET on various proteins indicate that some residues present specific mechanical properties that can be related to their biological activity, such as catalytic residues [38], residues controlling ligand migration within protein cavity networks [39,40], or residues located on the interdomain interfaces from multi-domain proteins [41,42]. In particular, an early work focusing on haemoproteins [43], and more specifically proteins presenting domains with globin or cytochrome c folds, showed that residues forming the folding nucleus in these two families could be identified via their remarkably high force constants within the protein structure.…”

Controlling Redox Enzyme Orientation at Planar Electrodes

“…Note that the enzyme orientation on the electrode is not only a key issue for interfacial electron transfer, but is also essential to facilitate the diffusion of substrates and products within the enzyme internal cavity network toward the catalytic site. Molecular Dynamics simulations on [NiFe] hydrogenases from A. aeolicus and Desulfovibrio fructosovorans have shown how these two homologous enzymes display their own specific internal tunnel network [31]. In the case of the membrane-bound A. aeolicus hydrogenase, all the substrate entry points are located on the same side Note that the enzyme orientation on the electrode is not only a key issue for interfacial electron transfer, but is also essential to facilitate the diffusion of substrates and products within the enzyme internal cavity network toward the catalytic site.…”

“…In the case of the membrane-bound A. aeolicus hydrogenase, all the substrate entry points are located on the same side Note that the enzyme orientation on the electrode is not only a key issue for interfacial electron transfer, but is also essential to facilitate the diffusion of substrates and products within the enzyme internal cavity network toward the catalytic site. Molecular Dynamics simulations on [NiFe] hydrogenases from A. aeolicus and Desulfovibrio fructosovorans have shown how these two homologous enzymes display their own specific internal tunnel network [31]. In the case of the Catalysts 2018, 8,192 7 of 38 membrane-bound A. aeolicus hydrogenase, all the substrate entry points are located on the same side of the enzyme's surface.…”

“…Apart from fast intramolecular ET achieved thanks to electronics relays within the enzyme, ET between physiological partners in a given chain, or between the substrate and the enzyme, is facilitated by specific recognition surfaces involving both hydrophobic and hydrophilic moieties [43]. As illustrations, the third, surface-exposed FeS cluster in the [NiFe]-Hases from Desulfovibrio is surrounded by an acidic patch composed of glutamic amino acid residues, which recognize a lysine-rich environment of the interacting heme of the physiological partner, a cytochrome c 3 [31,44]. This electrostatic interaction allows both partners to come to a close distance which then induces fast intermolecular ET.…”