Purification, characterization, and pituitary regulation of the sex-specific cytochrome P-450 15β-hydroxylase from liver microsomes of untreated female rats

C. MacGeoch, E. T. Morgan, J. Halpert, J. A. Gustafsson

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96 Scopus citations

Abstract

A sex-differentiated form of cytochrome P-450 has been purified to electrophoretic homogeneity from liver microsomes of untreated female rats. The purified preparation contained 17 nmol of P-450/mg of protein and had a minimum molecular weight of 50,000. The final preparation was active in the hydroxylation of 5α-[3H]androstane-3α,17β-diol 3,17-disulfate in the 15β position, with a turnover number of 2.6 nmol/min/nmol P-450 and is designated 'P-450 15β' on the basis of this activity. Cytochrome P-450 15β is isolated essentially in the low-spin state and has a CO-reduced difference spectral maximum at 449 nm. Comparison of the female protein with the corresponding P-450 fraction from male rats revealed an absence of the 15β band in the male electrophoretic profile. Specific antibodies to isozyme 15β were used with a Western blot technique to demonstrate the virtual absence of the protein in male microsomes. This method was also used to demonstrate that hypophysectomy of female rats resulted in undetectable levels of the 15β-hydroxylase, while continuous infusion of growth hormone to normal male animals increased the 15β-hydroxylase level to that of female. P-450 15β is proposed to be the enzyme responsible for the predominance of 15β-hydroxylated steroid sulfate metabolites in excreta of female rats, and their absence in males. The same purification procedure for female rat liver microsomes also yielded another purified cytochrome P-450 characterized by a minimum molecular weight of 52,500, termed P-450 DEa, which was inefficient in the 15β-hydroxylation of 5α-[3H]androstane-3α,17β-diol 3,17-disulfate. No evidence was obtained that this form is sexually differentiated.

Original languageEnglish (US)
Pages (from-to)15433-15439
Number of pages7
JournalJournal of Biological Chemistry
Volume259
Issue number24
StatePublished - 1984

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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