Kelly R, Miller S M, Lai M H, Kirsch D R
Squibb Institute for Medical Research, Department of Microbial Biochemistry and Genetics, Princeton, NJ 08543-4000.
Gene. 1990 Mar 15;87(2):177-83. doi: 10.1016/0378-1119(90)90299-7.
2,3-Oxidosqualene (OS) cyclase (OSC) catalyzes the conversion of OS to lanosterol, an essential step in the biosynthesis of sterols. The Candida albicans gene (ERG7) encoding OSC was cloned by complementation of a Saccharomyces cerevisiae OSC mutant (erg7). Two different Erg+ clones were isolated that contain a common overlapping region. The minimum region required for complementation was determined to be approx. 3.2 kb and a single 2.7-kb ERG7 transcript was detected. The cloned Candida ERG7 DNA complemented an additional nonconditional erg7 allele and a temperature-sensitive erg7 mutation. OSC activity was restored in the mutants as determined by [14C]acetate incorporation in vivo as well as incorporation in vitro in cell-free extracts using either [14C]isopentenyl pyrophosphate or [3H]OS as substrate. The level of OSC produced from expression of a single copy of the Candida ERG7 sequence was sufficient to allow growth of the S. cerevisiae erg7 mutants in the absence of exogenous ergosterol. These data support the contention that the Candida ERG7 sequence is the structural gene for OSC.
2,3-氧化角鲨烯（OS）环化酶（OSC）催化OS转化为羊毛甾醇，这是甾醇生物合成中的关键步骤。通过互补酿酒酵母OSC突变体（erg7）克隆了白色念珠菌编码OSC的基因（ERG7）。分离出两个不同的Erg +克隆，它们含有一个共同的重叠区域。互补所需的最小区域约为3.2 kb，并检测到一个单一的2.7 kb ERG7转录本。克隆的白色念珠菌ERG7 DNA补充了另一个非条件性erg7等位基因和一个温度敏感的erg7突变。通过体内[14C]乙酸掺入以及使用[14C]异戊烯基焦磷酸或[3H]OS作为底物在无细胞提取物中的体外掺入确定，突变体中的OSC活性得以恢复。由白色念珠菌ERG7序列单拷贝表达产生的OSC水平足以使酿酒酵母erg7突变体在没有外源性麦角固醇的情况下生长。这些数据支持白色念珠菌ERG7序列是OSC结构基因的论点。
