Electrochemistry of cytochrome P450 enzyme on nanoparticle-containing membrane-coated electrode and its applications for drug sensing

In the current study, we describe an improved system to study the two-electron delivery reaction pathway of cytochrome P450, family 2, subfamily B, polypeptide 6 (CYP2136) in vitro. In particular, a biocompatible film containing colloidal gold nanoparticles and chitosan was used to encapsulate CYP2136 on an electrode. The electrocatalytic behaviors of CYP2136 toward common drugs in the absence of NADHP-cytochrome P450 reductase as electron donor were studied. In an anaerobic solution, direct and reversible electron transfer between the electroactive heme center of CYP2B6 and the electrode was observed with a formal potential of -0.454 +/- 0.006 V at pH 7.4. In an air-saturated solution, an increase in the bioelectrocatalytic reduction current was observed after drug addition. The bioelectrocatalytic products were analyzed using high-performance liquid chromatography (HPLC) and electrospray ionization-mass spectrometry (ESI-MS). Both results confirmed that C-hydroxylation and heteroatom release were the main pathways for CYP2B6-mediated drug oxidation, similar to what occurred in vivo. The use of immobilized proteins in nanoparticle-containing films in drug biosensing was also demonstrated. (c) 2007 Elsevier Inc. All rights reserved.