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Orthogonal Translation Meets Electron Transfer: In Vivo Labeling of Cytochrome c for Probing Local Electric Fields
Author(s) -
Völler Jan,
Biava Hernan,
Koksch Beate,
Hildebrandt Peter,
Budisa Nediljko
Publication year - 2015
Publication title -
chembiochem
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.05
H-Index - 126
eISSN - 1439-7633
pISSN - 1439-4227
DOI - 10.1002/cbic.201500022
Subject(s) - heme , cytochrome c , electron transfer , redox , chemistry , hemeprotein , cytochrome , electron transport chain , cyclic voltammetry , biophysics , electrostatics , stereochemistry , electrochemistry , biochemistry , photochemistry , electrode , biology , organic chemistry , mitochondrion , enzyme
Cytochrome c (cyt c ), a redox protein involved in diverse fundamental biological processes, is among the most traditional model proteins for analyzing biological electron transfer and protein dynamics both in solution and at membranes. Studying the role of electric fields in energy transduction mediated by cyt c relies upon appropriate reporter groups. Up to now these had to be introduced into cyt c by in vitro chemical modification. Here, we have overcome this restriction by incorporating the noncanonical amino acid p ‐cyanophenylalanine ( p CNF) into cyt c in vivo. UV and CD spectroscopy indicate preservation of the overall protein fold, stability, and heme coordination, whereas a small shift of the redox potential was observed by cyclic voltammetry. The C≡N stretching mode of the incorporated p CNF detected in the IR spectra reveals a surprising difference, which is related to the oxidation state of the heme iron, thus indicating high sensitivity to changes in the electrostatics of cyt c .