Demonstration of a cytochrome P-450-dependent steroid 15β-hydroxylase in Bacillus megaterium

Anders Berg; Kjell Carlström; Jan Åke Gustafsson; Magnus Ingelman-Sundberg

doi:10.1016/0006-291X(75)90517-3

Demonstration of a cytochrome P-450-dependent steroid 15β-hydroxylase in Bacillus megaterium

Anders Berg, Kjell Carlström, Jan Åke Gustafsson, Magnus Ingelman-Sundberg

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Abstract

The steroid 15β-hydroxylase system of Bacillus megaterium was obtained in a cell-free preparation through sonication. The strictly NADPH-dependent 15β-hydroxylase activity, measured using progesterone as substrate, was inhibited by carbon monoxide, SKF 525-A, imidazole and metyrapone, indicating that the reaction is cytochrome P-450-dependent. A 40-fold purification of cytochrome P-450 in cell-free extracts was obtained by chromatography on DEAE-cellulose yielding a concentration of 0.32 nmoles of cytochrome P-450 per mg of protein. This partially purified cytochrome P-450 preparation catalyzed 15β- and 15α-hydroxylation of progesterone in the presence of NaIO₄ or NaClO₂ but not in the presence of NADPH or NADH.

Original language	English (US)
Pages (from-to)	1414-1423
Number of pages	10
Journal	Biochemical and Biophysical Research Communications
Volume	66
Issue number	4
DOIs	https://doi.org/10.1016/0006-291X(75)90517-3
State	Published - Oct 27 1975

ASJC Scopus subject areas

Biophysics
Biochemistry
Molecular Biology
Cell Biology

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title = "Demonstration of a cytochrome P-450-dependent steroid 15β-hydroxylase in Bacillus megaterium",

abstract = "The steroid 15β-hydroxylase system of Bacillus megaterium was obtained in a cell-free preparation through sonication. The strictly NADPH-dependent 15β-hydroxylase activity, measured using progesterone as substrate, was inhibited by carbon monoxide, SKF 525-A, imidazole and metyrapone, indicating that the reaction is cytochrome P-450-dependent. A 40-fold purification of cytochrome P-450 in cell-free extracts was obtained by chromatography on DEAE-cellulose yielding a concentration of 0.32 nmoles of cytochrome P-450 per mg of protein. This partially purified cytochrome P-450 preparation catalyzed 15β- and 15α-hydroxylation of progesterone in the presence of NaIO4 or NaClO2 but not in the presence of NADPH or NADH.",

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AU - Berg, Anders

AU - Carlström, Kjell

AU - Gustafsson, Jan Åke

AU - Ingelman-Sundberg, Magnus

PY - 1975/10/27

Y1 - 1975/10/27

N2 - The steroid 15β-hydroxylase system of Bacillus megaterium was obtained in a cell-free preparation through sonication. The strictly NADPH-dependent 15β-hydroxylase activity, measured using progesterone as substrate, was inhibited by carbon monoxide, SKF 525-A, imidazole and metyrapone, indicating that the reaction is cytochrome P-450-dependent. A 40-fold purification of cytochrome P-450 in cell-free extracts was obtained by chromatography on DEAE-cellulose yielding a concentration of 0.32 nmoles of cytochrome P-450 per mg of protein. This partially purified cytochrome P-450 preparation catalyzed 15β- and 15α-hydroxylation of progesterone in the presence of NaIO4 or NaClO2 but not in the presence of NADPH or NADH.

AB - The steroid 15β-hydroxylase system of Bacillus megaterium was obtained in a cell-free preparation through sonication. The strictly NADPH-dependent 15β-hydroxylase activity, measured using progesterone as substrate, was inhibited by carbon monoxide, SKF 525-A, imidazole and metyrapone, indicating that the reaction is cytochrome P-450-dependent. A 40-fold purification of cytochrome P-450 in cell-free extracts was obtained by chromatography on DEAE-cellulose yielding a concentration of 0.32 nmoles of cytochrome P-450 per mg of protein. This partially purified cytochrome P-450 preparation catalyzed 15β- and 15α-hydroxylation of progesterone in the presence of NaIO4 or NaClO2 but not in the presence of NADPH or NADH.

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Demonstration of a cytochrome P-450-dependent steroid 15β-hydroxylase in Bacillus megaterium

Abstract

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