Phospholipids of adrenal cortex mitochondria and the steroid hydroxylases: The lipid-environment of cytochrome P-450

doi:10.1016/S0006-291X(74)80361-X

Biochemical and Biophysical Research Communications

Volume 57, Issue 1, 15 March 1974, Pages 93-99

https://doi.org/10.1016/S0006-291X(74)80361-X Get rights and content

Summary

Adrenal cortex mitochondria have a high content of phospholipids with elevated ratios of phosphatidyl ethanolamine and phosphatidyl choline to diphosphatidyl glycerol, as compared with those of heart, liver, and kidney mitochondria. An extracted cytochrome P-450 preparation contained phosphatidyl ethanolamine and phosphatidyl choline as major components but no diphosphatidyl glycerol. After phospholipase-C pretreatment of the P-450 preparation, the reconstituted steroid 11 β-hydroxylase activity was inhibited, whereas the cholesterol desmolase activity was activated to a comparable extent.

References (17)

FleischerS. et al.
J. Lipid Res.
(1967)
CarafoliE. et al.
J. Biol. Chem.
(1971)
PéronF.G.
Biochem. Biophys. Acta
(1964)
PfeifferD.R. et al.
FEBS Letters
(1972)
CammerW. et al.
Arch. Biochem. Biophys.
(1967)
KimuraT. et al.
J. Biol. Chem.
(1967)
KimuraT. et al.
Anal. Biochem.
(1966)
ChuJ.W. et al.
J. Biol. Chem.
(1973)

There are more references available in the full text version of this article.

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Regulatory effect of copper on rat adrenal cytochrome P-450 and steroid metabolism
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: This study was supported by Research Grants from the National Institutes of Health (AM-12713, AM-13724, and AM-5384).

^*: Present address: Institute for Enzyme Research, University of Wisconsin, Madison, Wisconsin

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Phospholipids of adrenal cortex mitochondria and the steroid hydroxylases: The lipid-environment of cytochrome P-450

Summary

J. Lipid Res.

J. Biol. Chem.

Biochem. Biophys. Acta

FEBS Letters

Arch. Biochem. Biophys.

J. Biol. Chem.

Anal. Biochem.

J. Biol. Chem.