Figure 13 QCM-FIA data of oxidized cytochrome c at a lipid bilayer modified electrode containing no cytochrome c oxidase: (a) 10 µM cytochrome c in 0.10 M phosphate buffer; (b) 2.5 µM cytochrome c in 0.010 M phosphate buffer; (c) 5 µM cytochrome c in 0.010 M phosphate buffer; and (d) 10 µM cytochrome c in 0.010 M phosphate buffer. Buffer pH is 7.4 and the electrode area is 0.2 cm .

In evaluating the kinetics of this reaction under varied cytochrome c concentrations, a model has been assumed in which the rate-limiting step is not electron transfer from the cytochrome c oxidase to the electrode. The rate-limiting step is either electron transfer though the cytochrome c oxidase or between the cytochrome c in solution and the cytochrome c oxidase, depending on the experimental conditions. The biphasic kinetic response with increasing cytochrome c concentration is ascribed to the cytochrome c oxidase cycling between two conformations [77]. The two conformations form when electron transfer from protein in solution to the primary electron-accepting redox site of

the oxidase (Cua) occurs, and when electron transfer through the oxidase from Cua to the oxygen-binding site (Cub-heme a3) occurs. Oscillation between these two conformations also involves coupling of proton translocation with electron transfer through the

oxidase. As the cytochrome c concentration is increased above 10 µM, the rate of electron transfer from cytochrome c in solution to the electrode becomes limited by electron transfer through the cytochrome c oxidase. This is due to the rate of mediated electron transfer becoming limited by the rate of the conformation change of the oxidase from the electron injection conformation to the conformation required for electron transfer through the oxidase.