Direct electrochemistry of immobilized human cytochrome P450 2E1.
This communication reports the first electrochemical study of the human P450 2E1 either absorbed or covalently linked to different electrode surfaces. Glassy-carbon and gold electrodes gave reversible electrochemical signals of an active P450 2E1. Molecular modeling of the enzyme helped to rationalize the results. A monolayer coverage was obtained on gold modified with cystamine/maleimide that covalently linked surface accessible cysteines of P450 2E1. The midpoint potential measured for the oriented P450 2E1 was -177 +/- 5 mV comparable to that of the FeIII/FeII of other P450 enzymes. The observed electron-transfer rate for this electrode was 10 s-1. The turnover of the active enzyme was measured with the P450 2E1 specific substrate p-nitrophenol, resulting in a KM of 130 +/- 3 muM and the formation of 2.2 muM of the p-nitrocatechol product upon application of a -300 mV bias.
Fantuzzi
Andrea
A
Department of Biological Sciences, Imperial College, London, UK.
Fairhead
Michael
M
Gilardi
Gianfranco
G
D002244:Carbon; D002384:Catalysis; D019392:Cytochrome P-450 CYP2E1 / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004563:Electrochemistry / Q000379:methods*; D004566:Electrodes; D004800:Enzymes, Immobilized; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D006046:Gold; D006801:Humans; D008099:Liver / Q000201:enzymology; D009596:Nitrophenols / Q000737:chemistry; D010084:Oxidation-Reduction
D004800:Enzymes, Immobilized; D009596:Nitrophenols; D002244:Carbon; D006046:Gold; D019392:Cytochrome P-450 CYP2E1; C024836:4-nitrophenol