Direct electrochemistry of cytochrome P450 6A1 in mimic bio-membrane and its application for pesticides sensing

Abstract

The direct electrochemistry of house fly cytochrome P4506A1 (CYP6A1) confined in dioctadecyl dimethyl ammonium bromide (DDAB) film was achieved. The immobilized CYP6A1 displayed a pair of redox peaks with a formal potential of −0.36 mV in pH 7.0 O 2-free phosphate buffers at scan rate of 1 V s −1 and the direct electron transfer of CYP6A1 was characterized by voltammetry. The CYP6A1 in the DDAB film retained its bioactivity and could catalyze the reduction of dissolved oxygen. Upon addition of its substrate aldrin or heptachlor to the air-saturated solution, the reduction peak current of dissolved oxygen increased, which indicates the catalytic behavior of CYP6A1 to its substrates. By amperometry a calibration linear range was obtained to be 9.08 × 10 −6–4.54 × 10 −5 mol L −1 with a sensitivity of 80 μA mM −1 for aldrin or 8.91 × 10 −6–4.46 × 10 −5 mol L −1 with a sensitivity of 66 μA mM −1 for heptachlor. The apparent Michaelis–Menten constant for the electrocatalytic activity of CYP6A1 was found to be 7.468 × 10 −5 mol L −1 for aldrin and 4.316 × 10 −5 mol L −1 for heptachlor. The bioelectrocatalytic products were analysed using gas chromatography (GC) and electron ionization–mass spectrometry (EI–MS). The results confirmed that epoxidation was the main pathways of CYP6A1-mediated organochlorine pesticides oxidation.