2010
DOI: 10.1021/ac101072h
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Control of Human Cytochrome P450 2E1 Electrocatalytic Response as a Result of Unique Orientation on Gold Electrodes

Abstract: Oriented immobilization of human cytochrome P450 2E1 and its catalytic activity by direct electrochemistry was achieved by engineering two multisite mutants of P450 2E1: MUT261 (C268S-C480S-C488S) and MUT268 (C261S-C480S-C488S). Here, all the exposed cysteines are mutated into serines, with the exception of one (C261 for MUT261 and C268 for MUT268) that is able to link covalently to a modified gold electrode. The P450 2E1 wild type, as well as the two mutants, were immobilized onto gold electrodes using dithio… Show more

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Cited by 50 publications
(53 citation statements)
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“…Gold electrodes (11 mm × 5 mm, custom made by Arrandee, Germany) consisting of a vacuum evaporated thin gold film (thickness 200 nm) on borosilicate glass were annealed with a butane flame to obtain flat re-crystallized Au(111) terraces and were functionalized with dithio-bismaleimidoethane (DTME) as reported previously [12,[21][22][23]. This functionalization led to the formation of maleimide-terminated groups that subsequently interacted with exposed cysteine residues present on the surface of the M. fascicularis P450 2C20 (30 μM) resulting in covalent linkage of the protein to the Au surface.…”
Section: Immobilization Of P450 2c20 On Gold Electrodementioning
confidence: 99%
“…Gold electrodes (11 mm × 5 mm, custom made by Arrandee, Germany) consisting of a vacuum evaporated thin gold film (thickness 200 nm) on borosilicate glass were annealed with a butane flame to obtain flat re-crystallized Au(111) terraces and were functionalized with dithio-bismaleimidoethane (DTME) as reported previously [12,[21][22][23]. This functionalization led to the formation of maleimide-terminated groups that subsequently interacted with exposed cysteine residues present on the surface of the M. fascicularis P450 2C20 (30 μM) resulting in covalent linkage of the protein to the Au surface.…”
Section: Immobilization Of P450 2c20 On Gold Electrodementioning
confidence: 99%
“…Davis and Hill used scanning probe microscopy to characterize cytP450 cam monolayers on Au suitable for direct voltammetry 14. However, an unmet challenge in cyt P450 research has been to deliver electrons from electrode to CPR to cyt P450s to drive the natural catalytic cycle 15–18. Such “bioelectronic” initiation would enable electrochemical probing of the catalytic pathway, and simplify synthetic and biosensor applications by replacing electron donation from expensive NADPH to the enzyme.…”
Section: Introductionmentioning
confidence: 99%
“…[15][16][17][18][19][20][21][22][23][24] Particular interest has been focused on the improvement of the electronic coupling of the enzyme with the electrode surface and of its catalytic efficiency. In a fully coupled system reducing equivalents and molecular oxygen consumed are stoichiometrically equivalent to the product formed.…”
mentioning
confidence: 99%