Human sterol 14 alpha-demethylase activity is enhanced by the membrane-bound state of cytochrome b(5)

K. Venkateswarlu, Steven Lewis Kelly, Naheed N. Kaderbhai, David Christopher Lamb, Mustak A. Kaderbhai, Diane Elizabeth Kelly

Research output: Contribution to journalArticlepeer-review

19 Citations (SciVal)

Abstract

Human sterol 14α-demethylase (P45051; CYP51) catalyzes the oxidative removal of the C32 methyl group of dihydrolanosterol, an essential step in the cholesterol biosynthetic pathway. The reaction is dependent upon NADPH cytochrome P450 reductase (CPR) that donates the electrons for the catalytic cycle. Here we used a recombinant yeast CPR to investigate the abilities of four different forms of cytochrome b5 to support sterol demethylation activity of CYP51. The cytochrome b5 derivatives were genetically engineered forms of the native rat cytochrome b5 core-tail: the soluble globular b5 core (core), the core linked at its N-terminus with the secretory signal sequence of alkaline phosphatase (signal-core), and the signal sequence linked to the native b5 (signal-core-tail). The rat core-tail enzyme greatly stimulated sterol demethylation, whereas the signal-core-tail was only marginally active. In contrast, the core and signal-core constructs were completely inactive in stimulating the demethylation reaction. Additionally, cytochrome b5 enhanced sterol demethylation by more than threefold by accepting electrons from soluble yeast CPR and in its ability to reduce P450. We show that the nature of transient linkage between the hemoproteins and the redox partners is most likely brought about electrostatically, although productive interaction between cytochrome b5 and CYP51 is governed by the membrane-insertable hydrophobic region in the cytochrome b5 which in turn determines the correct spatial orientation of the core. This is the first report showing the stimulation of CYP51 by cytochrome b5.
Original languageEnglish
Pages (from-to)78-84
Number of pages7
JournalArchives of Biochemistry and Biophysics
Volume395
Issue number1
DOIs
Publication statusPublished - 01 Nov 2001

Keywords

  • cytochrome P450
  • cytochrome b5
  • sterol 14α-demethylase
  • steroidogenesis
  • electron transport

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