Dataset Information

Isolation and characterization of the gene encoding 2,3-oxidosqualene-lanosterol cyclase from Saccharomyces cerevisiae.

ABSTRACT: The ERG7 gene encoding oxidosqualene-lanosterol cyclase [(S)-2,3-epoxysqualene mutase (cyclizing, lanosterol forming), EC 5.4.99.7] from Saccharomyces cerevisiae has been cloned by genetic complementation of a cyclase-deficient erg7 strain. The DNA sequence of this gene has been determined and found to contain an open reading frame of 2196 nt (including stop codon) that encodes a predicted protein of 731 amino acids. The predicted molecular mass of the S. cerevisiae cyclase, 83.4 kDa, is similar to the predicted molecular masses of the oxidosqualene-lanosterol cyclase from Candida albicans and the oxidosqualene-cycloartenol cyclase from Arabidopsis thaliana, as well as to the molecular masses assigned to vertebrate oxidosqualene-lanosterol cyclases; however, it is substantially larger than the molecular mass assigned to purified S. cerevisiae cyclase. At the level of DNA and predicted amino acid sequences, the S. cerevisiae and C. albicans cyclases share 56% and 63% identity, respectively. Tryptophan and tyrosine residues are unusually abundant in the predicted amino acid sequences of (oxido)-squalene cyclases, leading to a hypothesis that electron-rich aromatic side chains from these residues are essential features of cyclase active sites.

SUBMITTER: Shi Z

PROVIDER: S-EPMC44401 | biostudies-other | 1994 Jul

REPOSITORIES: biostudies-other

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Isolation and characterization of the gene encoding 2,3-oxidosqualene-lanosterol cyclase from Saccharomyces cerevisiae.

Shi Z Z Buntel C J CJ Griffin J H JH

Proceedings of the National Academy of Sciences of the United States of America 19940701 15

The ERG7 gene encoding oxidosqualene-lanosterol cyclase [(S)-2,3-epoxysqualene mutase (cyclizing, lanosterol forming), EC 5.4.99.7] from Saccharomyces cerevisiae has been cloned by genetic complementation of a cyclase-deficient erg7 strain. The DNA sequence of this gene has been determined and found to contain an open reading frame of 2196 nt (including stop codon) that encodes a predicted protein of 731 amino acids. The predicted molecular mass of the S. cerevisiae cyclase, 83.4 kDa, is similar ...[more]

PMID: 8041797

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Project description:The effects of [4'-(6-allyl-methyl-amino-hexyloxy)-2'-fluoro-phenyl]-(4-bromophenyl)-methanone fumarate (Ro 48-8071), an inhibitor of 2,3-oxidosqualene:lanosterol cyclase (cyclase), were evaluated on CYP3A4 and CYP2B6 mRNA content in primary cultured human hepatocytes. In seven hepatocyte culture preparations, 24-h treatment with 3, 10, or 30 microM Ro 48-8071 produced median increases in CYP3A4 mRNA content that were 2.2-, 7.1-, and 8.5-fold greater than untreated control, respectively, and produced increases in CYP2B6 mRNA content that were 3.0-, 4.6-, and 3.4-fold greater than control, respectively. Increases in CYP3A4 immunoreactive protein content were also measured in Ro 48-8071-treated hepatocytes. To evaluate the effects of cyclase inhibitor treatments further, a pregnane X receptor (PXR)-responsive transactivation assay in HepG2 cells was used. Ro 48-8071, trans-N-(4-chlorobenzoyl)-N-methyl-(4-dimethylaminomethylphenyl)-cyclohexylamine (BIBX 79), and 3beta-(2-diethylaminoethoxy)androst-5-en-17-one HCl (U18666A) induced luciferase expression from a PXR-responsive reporter with EC(50)s of 0.113, 0.916, and 0.294 microM, respectively. Treatment of the HepG2 system with (E)N-ethyl-N-(6,6-dimethyl-2-hepten-4-ynyl)-3-[(3,3'-bithiophen-5-yl)methoxy]benzenemethanamine (NB-598), an inhibitor of squalene monooxygenase, at concentrations sufficient to achieve cholesterol biosynthesis inhibition significantly inhibited cyclase inhibitor-mediated, but not rifampicin-mediated, reporter induction. Direct treatment of the HepG2 system with 1 to 10 microM squalene 2,3:22,23-dioxide, but not squalene 2,3-oxide, significantly activated PXR-responsive reporter expression. Also, squalene 2,3:22,23-dioxide bound to human PXR in vitro with an IC(50) of 3.35 microM. These data indicate that cyclase inhibitors are capable of producing CYP3A4 and CYP2B6 induction in primary cultured human hepatocytes, and that an endogenous squalene metabolite is a conserved intracrine activator of PXR.

Dataset Information

Isolation and characterization of the gene encoding 2,3-oxidosqualene-lanosterol cyclase from Saccharomyces cerevisiae.

Publications

Isolation and characterization of the gene encoding 2,3-oxidosqualene-lanosterol cyclase from Saccharomyces cerevisiae.

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