UNMEDIATED HETEROGENEOUS ELECTRON-TRANSFER REACTION OF ASCORBATE OXIDASE AND LACCASE AT A GOLD ELECTRODE

The unmediated electrochemistry of two large Cu-containing proteins, a scorbate oxidase and laccase, was investigated by direct-current cycli c voltammetry. Rapid heterogeneous electron transfer was achieved in t he absence of promoters or mediators by trapping a small amount of pro tein within a solid, electrochemically inert, tributylmethyl phosphoni um chloride membrane coating a gold electrode. The problems typical of proteins in solution, such as adsorption on the electrode surface, we re avoided by this procedure. In anaerobic conditions, the cyclic volt ammograms, run at a scan rate of up to 200 mV/s, showed the electron t ransfer process to be quasi-reversible and diffusion-controlled. The p H-dependent redox potentials (+/-360 mV and +400 mV against a normal h ydrogen electrode at pH 7.0 for ascorbate oxidase and laccase respecti vely and +390 mV and +410 mV at pH 5.5) were similar to those of the f ree proteins. The same electrochemical behaviour was recorded for the type 2 Cu-depleted derivatives, which contain reduced type 3 Cu, where as the apoproteins were electrochemically inactive. Under aerobic cond itions the catalytic current intensity of holoprotein voltammograms in creased up to approx. 2-fold at a low scanning rate, with unchanged re dox potentials. The voltammograms of type 2 Cu-depleted proteins and o f apoproteins were unaffected by the presence of oxygen. This suggests that electron uptake at the electrode surface involves type 1 Cu and that only in the presence of oxygen is the intramolecular electron tra nsfer to other protein sites rapid enough to be observed. The analogy with available kinetic results is discussed.