Title

Reactive Sites and Course of Reduction in the Rieske Protein

Document Type

Article

Publication Date

6-2017

Abstract

Rieske proteins play an essential role in electron transfer in the bc1 complex. Rieske proteins contain a [2Fe–2S] cluster with one iron ligated by two histidines and the other iron ligated by two cysteines. All Rieske proteins have pH-dependent reduction potentials with the histidines ligating the cluster deprotonating in response to increases in pH. The addition of diethylpyrocarbonate (DEPC) modifies deprotonated histidines. The previous studies on the isolated Thermus thermophilus Rieske protein have used large excesses of DEPC, and this study examines what amino acids become modified under different molar equivalents of DEPC to protein. Increasing amounts of DEPC result in more modification, and higher pH values result in faster reaction. Upon modification, the protein also becomes reduced and ~6 equivalents of DEPC are needed for 50% of the reduction to occur. Which amino acids are modified first also points to the most reactive species on the protein. Mass spectrometry analysis shows that lysine 68 is the most reactive amino acid, followed by the ligating histidine 154 and two other surfaces lysines, 76 and 43. The modification of the ligating histidine at low numbers of DEPC equivalents and correlation with a similar number of equivalents needed to reduce the protein shows that this histidine can interact with neighboring groups, and these results can be extended to the protein within the bc1 complex, where interaction with neighboring residues or molecules may allow reduction to occur. These results may shed light on how Rieske transfers electrons and protons in the bc1 complex.

DOI

10.1007/s00775-017-1445-4

Publisher

Springer Nature

Publication Information

Journal of Biological Inorganic Chemistry

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