Coexpression of a human P450 (CYP3A4) and P450 reductase generates a highly functional monooxygenase system in Escherichia coli

Jennifer A. R. Blake; Mike Pritchard; Shaohong Ding; Graeme C. M. Smith; Brian Burchell; C.Roland Wolf; Thomas Friedberg

doi:10.1016/S0014-5793(96)01196-9

Coexpression of a human P450 (CYP3A4) and P450 reductase generates a highly functional monooxygenase system in Escherichia coli

Jennifer A. R. Blake, Mike Pritchard, Shaohong Ding, Graeme C. M. Smith, Brian Burchell, C.Roland Wolf, Thomas Friedberg

Research output: Contribution to journal › Article › peer-review

88 Citations (Scopus)

Abstract

The catalytic activities of recombinant cytochrome P450s expressed in E. coli have been impeded by the absence of endogenous P450 reductase. To solve this problem, we coexpressed P450 reductase with CYP3A4. Membranes from this strain contained 215 pmol P450/mg protein and a reductase activity of 1315 nmol cytochrome c reduced/min per mg. We detected 6ß-hydroxylation of testosterone and oxidation of nifedipine in vivo with turnover numbers of 15.2 and 17.3 min , respectively. These values compare favourably with those obtained using an optimally reconstituted system. Our data demonstrate that a catalytically efficient human P450 system can be generated in E. coli.

Original language	English
Pages (from-to)	210-214
Number of pages	5
Journal	FEBS Letters
Volume	397
Issue number	2-3
DOIs	https://doi.org/10.1016/S0014-5793(96)01196-9
Publication status	Published - 18 Nov 1996

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title = "Coexpression of a human P450 (CYP3A4) and P450 reductase generates a highly functional monooxygenase system in Escherichia coli",

abstract = "The catalytic activities of recombinant cytochrome P450s expressed in E. coli have been impeded by the absence of endogenous P450 reductase. To solve this problem, we coexpressed P450 reductase with CYP3A4. Membranes from this strain contained 215 pmol P450/mg protein and a reductase activity of 1315 nmol cytochrome c reduced/min per mg. We detected 6{\ss}-hydroxylation of testosterone and oxidation of nifedipine in vivo with turnover numbers of 15.2 and 17.3 min , respectively. These values compare favourably with those obtained using an optimally reconstituted system. Our data demonstrate that a catalytically efficient human P450 system can be generated in E. coli.",

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AU - Blake, Jennifer A. R.

AU - Pritchard, Mike

AU - Ding, Shaohong

AU - Smith, Graeme C. M.

AU - Burchell, Brian

AU - Wolf, C.Roland

AU - Friedberg, Thomas

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N2 - The catalytic activities of recombinant cytochrome P450s expressed in E. coli have been impeded by the absence of endogenous P450 reductase. To solve this problem, we coexpressed P450 reductase with CYP3A4. Membranes from this strain contained 215 pmol P450/mg protein and a reductase activity of 1315 nmol cytochrome c reduced/min per mg. We detected 6ß-hydroxylation of testosterone and oxidation of nifedipine in vivo with turnover numbers of 15.2 and 17.3 min , respectively. These values compare favourably with those obtained using an optimally reconstituted system. Our data demonstrate that a catalytically efficient human P450 system can be generated in E. coli.

AB - The catalytic activities of recombinant cytochrome P450s expressed in E. coli have been impeded by the absence of endogenous P450 reductase. To solve this problem, we coexpressed P450 reductase with CYP3A4. Membranes from this strain contained 215 pmol P450/mg protein and a reductase activity of 1315 nmol cytochrome c reduced/min per mg. We detected 6ß-hydroxylation of testosterone and oxidation of nifedipine in vivo with turnover numbers of 15.2 and 17.3 min , respectively. These values compare favourably with those obtained using an optimally reconstituted system. Our data demonstrate that a catalytically efficient human P450 system can be generated in E. coli.

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Coexpression of a human P450 (CYP3A4) and P450 reductase generates a highly functional monooxygenase system in Escherichia coli

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