An electrode surface is presented that enables the characterization of redox-active membrane enzymes in a native-like environment. An ubiquinol oxidase from Escherichia coli, cytochrome bo3 (cbo3), has been co-immobilized into tethered bilayer lipid membranes (tBLMs). The tBLM is formed on gold surfaces functionalized with cholesterol tethers which insert into the lower leaflet of the membrane. The planar membrane architecture is formed by self-assembly of proteoliposomes, and its structure is characterized by surface plasmon resonance (SPR), electrochemical impedance spectroscopy (EIS), and tapping-mode atomic force microscopy (TM-AFM). The functionality of cbo 3 is investigated by cyclic voltammetry (CV) and is confirmed by the catalytic reduction of oxygen. Interfacial electron transfer to cbo3 is mediated by the membrane-localized ubiquinol-8, the physiological electron donor of cbo3. Enzyme coverages observed with TM-AFM and CV coincide (2-8.5 fmol·cm-2), indicating that most-if not all-cbo 3 on the surface is catalytically active and thus retains its integrity during immobilization.
|Original language||English (US)|
|Number of pages||6|
|Journal||Journal of the American Chemical Society|
|State||Published - Feb 8 2006|
ASJC Scopus subject areas
- Colloid and Surface Chemistry