SERR Spectroelectrochemical Study of Cytochrome cd (1) Nitrite Reductase Co-Immobilized with Physiological Redox Partner Cytochrome c (552) on Biocompatible Metal Electrodes

Cytochrome cd1 nitrite reductases (cd (1)NiRs) catalyze the one-electron reduction of nitrite to nitric oxide. Due to their catalytic reaction, cd (1)NiRs are regarded as promising components for biosensing, bioremediation and biotechnological applications. Motivated by earlier findings that catalytic activity of cd (1)NiR from Marinobacter hydrocarbonoclasticus (Mhcd (1)) depends on the presence of its physiological redox partner, cytochrome c (552) (cyt c (552)), we show here a detailed surface enhanced resonance Raman characterization of Mhcd (1) and cyt c (552) attached to biocompatible electrodes in conditions which allow direct electron transfer between the conducting support and immobilized proteins. Mhcd (1) and cyt c552 are co-immobilized on silver electrodes coated with self-assembled monolayers (SAMs) and the electrocatalytic activity of Ag // SAM // Mhcd (1) // cyt c (552) and Ag // SAM // cyt c (552) // Mhcd (1) constructs is tested in the presence of nitrite. Simultaneous evaluation of structural and thermodynamic properties of the immobilized proteins reveals that cyt c (552) retains its native properties, while the redox potential of apparently intact Mhcd (1) undergoes a ~150 mV negative shift upon adsorption. Neither of the immobilization strategies results in an active Mhcd (1), reinforcing the idea that subtle and very specific interactions between Mhcd (1) and cyt c (552) govern efficient intermolecular electron transfer and catalytic activity of Mhcd (1).