A mediated thin-layer voltammetry method for the study of redox protein electrochemistry.
A novel mediated thin-layer voltammetry technique that allows the rapid determination of midpoint potentials and electron transfer rate constants for small quantities of redox active proteins is described. Thin-layer voltammograms simulated for an electrolyte containing a redox active protein and an electron transfer mediator show that the rapid homogeneous electron exchange reaction between the protein and the mediator serves to mediate the charge transfer of the protein at the electrode, which does not take place in the absence of the mediator, and results in the observation of an apparently reversible redox couple. Both theoretical and experimental data are presented which suggest that the thin-layer voltammetry method will be generally applicable for the determination of protein redox potentials with the proper selection of mediators. Rate constants for the electron transfer between metalloproteins and mediators can be evaluated by comparing experimental voltammograms with theoretical data from simulations. The technique is demonstrated for the metalloproteins cytochrome c, ferredoxin, and the iron protein of nitrogenase.
Parker
V D
VD
Department of Chemistry and Biochemistry, Utah State University, Logan 84322-0300, USA. vparker@cc.usu.edu
Seefeldt
L C
LC
D000818:Animals; D016948:Azotobacter vinelandii / Q000378:metabolism; D003574:Cytochrome c Group / Q000737:chemistry / Q000378:metabolism; D004563:Electrochemistry / Q000295:instrumentation / Q000379:methods*; D004566:Electrodes; D004579:Electron Transport; D005288:Ferredoxins / Q000737:chemistry / Q000378:metabolism; D066298:In Vitro Techniques; D007700:Kinetics; D008667:Metalloproteins / Q000737:chemistry / Q000378:metabolism; D009591:Nitrogenase / Q000737:chemistry / Q000378:metabolism; D010084:Oxidation-Reduction; D010088:Oxidoreductases; D011506:Proteins / Q000737:chemistry* / Q000378:metabolism
D003574:Cytochrome c Group; D005288:Ferredoxins; D008667:Metalloproteins; D011506:Proteins; D010088:Oxidoreductases; D009591:Nitrogenase; C029130:nitrogenase reductase