Orthogonal Translation Meets Electron Transfer: In Vivo Labeling of Cytochrome c for Probing Local Electric Fields |
| |
| Authors: | Jan Völler Dr. Hernan Biava Prof. Beate Koksch Prof. Peter Hildebrandt Prof. Nediljko Budisa |
| |
| Affiliation: | 1. Institut für Chemie, Technische Universit?t Berlin, Müller‐Breslau‐Strasse 10, 10623 Berlin (Germany);2. Institut für Chemie und Biochemie, Freie Universit?t Berlin, Takustrasse 3, 14195 Berlin (Germany);3. Institut für Chemie, Technische Universit?t Berlin, Strasse des 17. Juni 135, 10623 Berlin (Germany) |
| |
| Abstract: | 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 (pCNF) 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 pCNF 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. |
| |
| Keywords: | cyanophenylalanine cytochromes infrared spectroscopy non‐canonical amino acids redox chemistry stop codon suppression |
|
|
