No abstract

Molekularer Lichtschalter: Am Ein‐/Ausschalten der Fluoreszenz des Proteins asFP595 ist eine trans‐cis‐Isomerisierung beteiligt (siehe Schema). Quantenmechanische und klassische Simulationen erhellen die spektroskopischen Eigenschaften von asFP595 und verschaffen einen genauen Einblick in den Photoschaltmechanismus. Der trans‐cis‐Konformationswechsel löst eine Protonentransferkaskade zwischen dem Chromophor und benachbarten Aminosäuren aus.

Molecular light‐switch: Off–on switching of the fluorescence of the protein asFP595 involves a trans–cis isomerization. Mixed quantum/classical simulations elucidate the spectroscopic properties of asFP595 and give detailed insights into the photoswitching mechanism. The conformational trans–cis switching triggers a proton‐transfer cascade between the chromophore and adjacent amino acids.

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.

This paper presents a theoretical analysis of the titration behavior of strongly interacting titratable residues in proteins. Strongly interacting titratable residues exist in many proteins such as for instance bacteriorhodopsin, cytochrome c oxidase, cytochrome bc(1), or the photosynthetic reaction center. Strong interaction between titratable groups can lead to irregular titration behavior. We analyze under which circumstances titration curves can become irregular. We demonstrate that conformational flexibility alone can not lead to irregular titration behavior. Strong interaction between titratable groups is a necessary, but not sufficient condition for irregular titration curves. In addition, the two interacting groups also need to titrate in the same pH-range. These two conditions together lead to irregular titration curves. The mutation of a single residue within a cluster of interacting titratable residues can influence the titration behavior of the other titratable residues in the cluster. We demonstrate this effect on a cluster of four interacting residues. This example underlines that mutational studies directed at identifying the role of a certain titratable residue in a cluster of interacting residues should always be accompanied by an analysis of the effect of the mutation on the titration behavior of the other residues.