Molecularly wiring of Cytochrome c with carboxylic acid functionalized hydroquinone on MWCNT surface and its bioelectrocatalytic reduction of H2O2 relevance to biomimetic electron-transport and redox signalling

Abstract

The electron-transport chain that involves a proton-coupled electron-transfer (ET) reaction between hydroquinone (H2Q) and Cytochrome c (Cyt c) in the acid pool is one of the key processes of the respiratory complex 1 in cytoplasmic membrane of the bacterium and mitochondrial complexes, complex III (cytochrome bc1 or ubiquinol:cytochrome c oxidoreductase) of photosynthesis systems. In the literature, several spectroscopic techniques in association with electron-transfer inhibitors have been reported for this purpose. Till now, there is no straight forward voltammetric proof to reveal the mechanistic feature of the direct ET process. In the electrochemistry point of view, it is a challenging task to prepare a chemically modified bio-electrode composed of H2Q and Cytc redox systems with proper ET channels between them and to probe the mixed-potential reaction by direct voltammetric technique. Herein, we report a carboxylic acid functionalized hydroquinone (2,5-dihydroxy benzoic acid; H2Q-COOH) immobilized multiwalled carbon nanotube modified glassy carbon electrode (GCE/MWCNT@H2Q-COOH), prepared by in-situ electrochemical oxidation of 2-hydroxy benzoic acid (Ph-COOH), as a molecular wiring system for unfolded Cytc protein and further to study the direct voltammetric ET reaction using H2O2 as a specific probe in pH 2 KCl-HCl medium. The formation of surface-confined H2Q-COOH was confirmed by physicochemical and spectroscopic characterization techniques including IR, Raman, UV-Vis, Thin-layer chromatography, NMR and GC-MS. The GCE/MWCNT@H2Q-COOH showed well-defined redox peaks corresponding to consecutive one-electron-transfer reactions of H2Q-COOH/•QH-COOH (Eo'= 0.410 V vs Ag/AgCl, A1/C1) and •QH-COOH/Q (Eo'= 0.750 V vs Ag/AgCl, A2/C2), similar to one occurs with a membrane-bound ubiquinone and plastoquinone systems in the biological systems. The unfolded Cytc modified electrode, GCE/MWCNT@H2Q-COOH@Cytc prepared by chemical modification technique, has exhibited a specific and selective bioelectrocatalytic reduction current signal in between the redox potentials of QH2/•QH and Cytc in pH 2 KCl-HCl biomimicking the biological electron-transport chain reaction. In further amperometric techniques have been used to successfully demonstrate the ET reaction.