Direct electrochemistry of cytochrome P450 reductases in surfactant and polyion films

NADPH-cytochrome P450 reductase (CPR) serves as electron donor for cytochrome P450 catalyzed monooxygenase reactions utilizing flavin mononucleotide (FMN) and flavin adenine dinucleotide (FAD) as electron transfer cofactors. Here, stable films of human and rabbit CPRs with didodecyldimethylammonium bromide (DDAB), dimyristoylphosphatidyl choline (DMPC), and poly(diallyidimethylammonium) (PDDA) were made on pyrolytic graphite (PG) electrodes for comparative structural and electrochemical studies. CD and UV-VIS absorbance spectra suggested that near native CPR conformation is retained in PDDA films, and some conformational changes occur in DMPC or DDAB films. Cyclic voltammetry of these films gave quasireversible pairs of peaks at average formal potential -0.246 +/- 0.008 V vs. NHE. In human CPR-DDAB (H-CPR-DDAB), a second pair of peaks at +0.317 V vs. NHE was found that depended strongly on identity of buffer and salt. Excepting H-CPR in DDAB, films showed similar voltammetry, formal potentials, and k(s) values. While CPR-PDDA films had near native CPR structures, electrochemical parameters did not differ significantly from CPR-DMPC films. The relative independence of film voltammetry from the influence of film materials for CPRs is in contrast with heme iron proteins that, while retaining near native structures, have formal potentials that depend significantly on identity of the film material.