ABTS oxidation reaction as a model of cytochrome c-driven electron transfer

Cytochrome c, as an electron carrier within the mitochondria, plays a crucial role in the electron transport chain. To meet the demand for rapid methods that assess the electron transport properties of cytochrome c, we used the electron donor 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulphonate) (ABTS) as a substrate and suitable spectrophotometric reporter of cytochrome c peroxidase-like activity. ABTS and cytochrome c from bovine were purchased from Sigma-Aldrich Inc. The time course of the cytochrome c-driven ABTS oxidation reaction was studied using H2O2 as a second substrate. It was demonstrated that CytC addition is a prerequisite for the transfer of electrons from ABTS to H2O2. The reaction kinetic analysis with determination of Vmax, Km, kcat, and kcat/Km values for both substrates was performed. Our results demonstrate that the cytochrome c-catalyzed ABTS oxidation reaction can be effectively employed as a model for studying the functional role of cytochrome c in various conditions. Keywords: ABTS reaction, cytochrome c, electron transport rate, Michaelis-Menten kinetics parameters, peroxidase activity