Klingen Astrid R, Ullmann G Matthias
Department of Structural Biology/Bioinformatics, Bayreuth University, Universitätsstrasse 30, BGI, D-95447 Bayreuth, Germany.
Biochemistry. 2004 Oct 5;43(39):12383-9. doi: 10.1021/bi0488606.
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.
铁硫蛋白携带一个氧化还原活性铁硫簇,该簇由两个组氨酸和两个半胱氨酸侧链结合。铁硫蛋白的还原电位取决于pH值。这种pH依赖性可以用两个pK(a)值来描述,这两个值已被指定给两个与铁配位的组氨酸。铁硫蛋白通常分为两大类:来自喹啉氧化细胞色素bc复合物的铁硫蛋白,其中配体组氨酸在生理pH范围内滴定;以及细菌铁氧还蛋白铁硫蛋白,其中配体组氨酸在生理pH下质子化。在本文介绍的研究中,我们使用密度泛函理论/连续介质静电学相结合的方法计算了簇配体组氨酸的pK(a)值。可以重现bc型和铁氧还蛋白铁硫蛋白的实验pK(a)值。我们可以识别这两种蛋白质在功能上的重要差异:bc型铁硫蛋白中存在的朝向簇的氢键,以及铁氧还蛋白铁硫蛋白中存在的带负电荷的残基。我们通过在计算中对蛋白质进行突变消除了这些差异。突变蛋白中的铁硫中心具有非常相似的pK(a)值。因此,我们得出结论,所研究的结构差异是蛋白质不同pH滴定行为的主要原因。有趣的是,铁氧还蛋白铁硫蛋白中通过中和负电荷引起的偏移大于bc型铁硫蛋白中通过去除朝向簇的氢键引起的偏移。