Extensive quantum chemical DFT calculations were performed on the high-resolution (1.9 Å) crystal structure of photosystem II in order to determine the protonation pattern and the oxidation states of the oxygen-evolving Mn cluster. First, our data suggest that the experimental structure is not in the S(1)-state. Second, a rather complete set of possible protonation patterns is studied, resulting in very few alternative protonation patterns whose relevance is discussed. Finally, we show that the experimental structure is a mixture of states containing highly reduced forms, with the largest contribution (almost 60%) from the S(-3)-state, Mn(II,II,III,III).

Density functional theory (DFT) was combined with solution of the Poisson equation for continuum dielectric media to compute accurate redox potentials for several mononuclear transition metal complexes (TMCs) involving iron, manganese, and nickel. Progress was achieved by altering the B3LYP DFT functional (B4(XQ3)LYP-approach) and supplementing it with an empirical correction term G(X) having three additional adjustable parameters, which is applied after the quantum-chemical DFT computations. This method was used to compute 58 redox potentials of 48 different TMCs involving different pairs of redox states solvated in both protic and aprotic solvents. For the 58 redox potentials the root mean square deviation (RMSD) from experimental values is 65 mV. The reliability of the present approach is also supported by the observation that the energetic order of the spin multiplicities of the electronic ground states is fulfilled for all studied TMCs, if the influence from the solvent is considered as well.

Cytochrome c oxidase (CcO) is a central enzyme in aerobic life catalyzing the conversion of molecular oxygen to water and utilizing the chemical energy to pump protons and establish an electrochemical gradient. Despite intense research, it is not understood how CcO achieves unidirectional proton transport and avoids short circuiting the proton pump. Within this work, we analyzed the potential role of Glu286 as a proton valve. We performed unconstrained MD simulations of CcO with an explicit membrane for up to 80 ns. Those MD simulations revealed that deprotonated Glu286 (Glu286-) is repelled by the negatively charged propionic acid PRD of heme a3. Thus, it destabilizes a potential linear chain of waters in the hydrophobic cavity connecting Glu286 with PRD and the binuclear center (BNC). Conversely, protonated Glu286 (Glu286H) may remain in an upward position (oriented toward PRD) and can stabilize the connecting linear water chain in the hydrophobic cavity. We calculated the pKa of Glu286 under physiological conditions to be above 12, but this value decreases to about 9 under increased water accessibility of Glu286. The latter value is in accordance with experimental measurements. In the time course of MD simulation, we also observed conformations where Glu286 bridges between water molecules located on both sides (the D channel being connected to the N side and the hydrophobic cavity), which might lead to proton backflow.

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Density functional theory computations of heme with ionized propionic acid groups, axially coordinated with two imidazoles, were performed for different mutual orientations of the imidazole planes. Environmental influences from water or protein were considered with a continuum dielectric medium by solving the Poisson equation. In vacuum, optimized geometries yielded imidazole-heme conformations where the NH groups of imidazoles are oriented toward the heme propionic groups in agreement with data from crystal structures of heme proteins. Conformational free-energy dependencies of the mutual orientation of axially ligated imidazoles calculated in protein (epsilon=10) and water (epsilon=80) environments confirmed the vacuum results, albeit the energy difference between the preferred and the 180 degrees opposite orientations of the imidazole ligand decreased from 3.84 kcal/mol in vacuum to 2.35 and 2.40 kcal/mol in protein and water, respectively. Two main factors determine the imidazole orientation: (1) the direct intramolecular electrostatic interactions of propionic groups with the polar NH groups of imidazole and (2) the electrostatic interaction of the total dipole moment of the imidazole-heme complex with the reaction field. In vacuum, only the first type of interaction is present, while in a dielectric medium the latter effect becomes competitive at high dielectric constant, resulting in a decrease of the orientational preference. Interestingly, the orientational preference of the imidazole axially ligated to heme becomes even more pronounced, if the negatively charged propionates are neutralized by counter charges that mimic salt bridges or protonation of the propionates.