Bard M, Bruner D A, Pierson C A, Lees N D, Biermann B, Frye L, Koegel C, Barbuch R
Department of Biology, Indiana University-Purdue University at Indianapolis, IN 46202, USA.
Proc Natl Acad Sci U S A. 1996 Jan 9;93(1):186-90. doi: 10.1073/pnas.93.1.186.
We have cloned the Saccharomyces cerevisiae C-4 sterol methyl oxidase ERG25 gene. The sterol methyl oxidase performs the first of three enzymic steps required to remove the two C-4 methyl groups leading to cholesterol (animal), ergosterol (fungal), and stigmasterol (plant) biosynthesis. An ergosterol auxotroph, erg25, which fails to demethylate and concomitantly accumulates 4,4-dimethylzy-mosterol, was isolated after mutagenesis. A complementing clone consisting of a 1.35-kb Dra I fragment encoded a 309-amino acid polypeptide (calculated molecular mass, 36.48 kDa). The amino acid sequence shows a C-terminal endoplasmic reticulum retrieval signal KKXX and three histidine-rich clusters found in eukaryotic membrane desaturases and in a bacterial alkane hydroxylase and xylene monooxygenase. The sterol profile of an ERG25 disruptant was consistent with the erg25 allele obtained by mutagenesis.
我们克隆了酿酒酵母C-4甾醇甲基氧化酶ERG25基因。甾醇甲基氧化酶在去除两个C-4甲基的三个酶促步骤中执行第一步，这两个C-4甲基是胆固醇（动物）、麦角固醇（真菌）和豆甾醇（植物）生物合成所必需的。在诱变后分离出一种麦角固醇营养缺陷型菌株erg25，它不能进行去甲基化并同时积累4,4-二甲基酵母甾醇。一个由1.35 kb的Dra I片段组成的互补克隆编码一个309个氨基酸的多肽（计算分子量为36.48 kDa）。氨基酸序列显示其C末端有内质网回收信号KKXX，以及在真核膜去饱和酶、细菌烷烃羟化酶和二甲苯单加氧酶中发现的三个富含组氨酸的簇。ERG25破坏株的甾醇谱与通过诱变获得的erg25等位基因一致。
