A combined fluorescence spectroscopic and electrochemical approach for the study of thioredoxins.
A new way to study the electrochemical properties of proteins by coupling front-face fluorescence spectroscopy with an optically transparent thin-layer electrochemical cell is presented. First, the approach was examined on the basis of the redox-dependent conformational changes in tryptophans in cytochrome c, and its redox potential was successfully determined. Second, an electrochemically induced fluorescence analysis of periplasmic thiol-disulfide oxidoreductases SoxS and SoxW was performed. SoxS is essential for maintaining chemotrophic sulfur oxidation of Paracoccus pantotrophus active in vivo, while SoxW is not essential. According to the potentiometric redox titration of tryptophan fluorescence, the midpoint potential of SoxS was -342 ± 8 mV versus the standard hydrogen electrode (SHE') and that of SoxW was -256 ± 10 mV versus the SHE'. The fluorescence properties of the thioredoxins are presented and discussed together with the intrinsic fluorescence contribution of the tyrosines.
Laboratoire de Spectroscopie Vibrationnelle et Electrochimie des Biomolécules, UMR 7177, Institut de Chimie, CNRS-Université de Strasbourg, 1 rue Blaise Pascal, 67070 Strasbourg, France.
Mariana
M
Voicescu
Dagmar
D
Rother
Frank
F
Bardischewsky
Cornelius G
CG
Friedrich
Petra
P
Hellwig
D000595:Amino Acid Sequence; D000818:Animals; D001426:Bacterial Proteins / Q000737:chemistry*; D045304:Cytochromes c / Q000737:chemistry*; D055664:Electrochemical Techniques / Q000295:instrumentation / Q000379:methods; D004867:Equipment Design; D006736:Horses; D008969:Molecular Sequence Data; D010084:Oxidation-Reduction; D010088:Oxidoreductases / Q000737:chemistry*; D042084:Paracoccus pantotrophus / Q000737:chemistry* / Q000201:enzymology; D013050:Spectrometry, Fluorescence / Q000295:instrumentation* / Q000379:methods; D013879:Thioredoxins / Q000737:chemistry*
D001426:Bacterial Proteins; D013879:Thioredoxins; D045304:Cytochromes c; D010088:Oxidoreductases