Negatively Charged Residues and Hydrogen Bonds Tune the Ligand Histidine pK_a Values of Rieske Iron−Sulfur Proteins

Abstract

Rieske proteins carry a redox-active iron-sulfur cluster, which is bound by two histidine and two cysteine side chains. The reduction potential of Rieske proteins depends on pH. This pH dependence can be described by two pK(a) values, which have been assigned to the two iron-coordinating histidines. Rieske proteins are commonly grouped into two major classes: Rieske proteins from quinol-oxidizing cytochrome bc complexes, in which the ligand histidines titrate in the physiological pH range, and bacterial ferredoxin Rieske proteins, in which the ligand histidines are protonated at physiological pH. In the study presented here, we have calculated pK(a) values of the cluster ligand histidines using a combined density functional theory/continuum electrostatics approach. Experimental pK(a) values for a bc-type and a ferredoxin Rieske protein could be reproduced. We could identify functionally important differences between the two proteins: hydrogen bonds toward the cluster, which are present in bc-type Rieske proteins, and negatively charged residues, which are present in ferredoxin Rieske proteins. We removed these differences by mutating the proteins in our calculations. The Rieske centers in the mutated proteins have very similar pK(a) values. We thus conclude that the studied structural differences are the main reason for the different pH-titration behavior of the proteins. Interestingly, the shift caused by neutralizing the negative charges in ferredoxin Rieske proteins is larger than the shift caused by removing the hydrogen bonds toward the cluster in bc-type Rieske proteins.

Reduction Potentials and Acidity Constants of Mn Superoxide Dismutase Calculated by QM/MM Free‐Energy Methods

“…However, several attempts have been made, especially of metal-bound water molecules, using the LD and PB methods. [18,24,27,31,35,36,40,43,44] The situation is analogous for reduction potentials, although the number of publications is smaller. Several scientists have tried to estimate the reduction potential of various groups in solution and in proteins, involving both metal sites and other redox-active cofactors.…”

Catalytic Cycle of Multicopper Oxidases Studied by Combined Quantum- and Molecular-Mechanical Free-Energy Perturbation Methods

“…However, several attempts have been made, especially for metal-bound water molecules, using the LD and PB methods. 32,34,[52][53][54] In this work, we study electron-and proton transfer reactions in the reaction cycle of the MCOs with the help of FEP calculations at the combined QM and MM (QM/MM) level. By estimating redox potentials, acidity constants, isomerisation free energies, as well as O 2 and water-binding free energies of putative intermediates, we obtain information about the most likely steps in the reaction mechanism, especially for the regeneration of the reduced state from the NI.…”

Modifications of Protein Environment of the [2Fe-2S] Cluster of the bc1 Complex

“…Consideration of the role of changes in pK introduced an additional constraint on the mechanism (15). In the work of Klingen and Ullmann (21), the pK a values of Rieske iron-sulfur clusters of bc-type and ferredoxin-type were calculated using density functional theory/electrostatic approaches based on the protein environment, and they suggested that the high E m and low pK a values in the former were largely determined by hydrogen bonding from these and other groups around the cluster.…”