Expression of a rat liver microsomal cytochrome P-450 catalyzing testosterone 16α-hydroxylation in Saccharomyces cerevisiae: Vitamin D3 25-hydroxylase and testosterone 16α-hydroxylase are distinct forms of cytochrome P-450

Shin Ichi Hayashi, Ken Ichirou Morohashi, Hidefumi Yoshioka, Kyuichiro Okuda, Tsuneo Omura

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19 Citations (Scopus)

Abstract

Rat cytochrome P-450(M-1) cDNA was expressed in Saccharomyces cerevisiae TD1 cells by using a yeast-Escherichia coli shuttle vector consisting of P-450(M-1) cDNA, yeast alcohol dehydrogenase promoter and yeast cytochrome c terminator. The yeast cells synthesized up to 2 × 105 molecules of P-450(M-1) per cell. The microsomal fraction prepared from the transformed cells contained 0.1 nmol of cytochrome P-450 per mg of protein. The expressed cytochrome P-450 catalyzed 16α- and 2α-hydroxylations of testosterone in accordance with the catalytic activity of P-450(M-1), but did not hydroxylate vitamin D3 or lα-hydroxycholecalciferol at the 25 position. The expressed cytochrome P-450 also catalyzed the oxidation of several drugs and did not show 25-hydroxylation activity toward 5β-cholestane-3α, 7α, 12α-triol. However, it cross-reacted with the polyclonal and monoclonal antibodies elicited against purified P-450cc25 which catalyzed the 25-hydroxylation of vitamin D3. These results indicated that P-450(M-1) cDNA coded the 2α- and 16α-hydroxylase of testosterone, and that these two positions of testosterone are hydroxylated by a single form of cytochrome P-450. Vitamin D3 25-hydroxylase and testosterone 16α-and 2α-hydroxylase are different gene products, although these two hydroxylase activities are immunochemically indistinguishable.

Original languageEnglish
Pages (from-to)858-862
Number of pages5
JournalJournal of biochemistry
Volume103
Issue number5
DOIs
Publication statusPublished - May 1988

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Molecular Biology

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