Chang S C, Heacock P N, Mileykovskaya E, Voelker D R, Dowhan W
Department of Biochemistry and Molecular Biology, University of Texas Medical School, Houston, Texas 77225, USA.
J Biol Chem. 1998 Jun 12;273(24):14933-41. doi: 10.1074/jbc.273.24.14933.
In eukaryotic cells, cardiolipin (CL) synthase catalyzes the final step in the synthesis of CL from phosphatidylglycerol and CDP-diacylglycerol. CL and its synthesis are localized predominantly to the mitochondrial inner membrane, and CL is generally thought to be an essential component of many mitochondrial processes. By using homology searches for genes potentially encoding phospholipid biosynthetic enzymes, we have cloned the gene (CLS1) encoding CL synthase in Saccharomyces cerevisiae. Overexpression of the CLS1 gene under its endogenous promoter or the inducible GAL1 promoter in yeast and expression of CLS1 in baculovirus-infected insect cells resulted in elevated CL synthase activity. Disruption of the CLS1 gene in a haploid yeast strain resulted in the loss of CL synthase activity, no detectable CL, a 5-fold elevation in phosphatidylglycerol levels, and lack of staining of mitochondria by a dye with high affinity for CL. The cls1::TRP1 null mutant grew on both fermentable and non-fermentable carbon sources but more poorly on the latter. The level and activity of cytochrome c oxidase was normal, and a dye whose accumulation is dependent on membrane proton electrochemical potential effectively stained the mitochondria. These results definitively identify the gene encoding the CL synthase of yeast.
在真核细胞中，心磷脂（CL）合酶催化由磷脂酰甘油和CDP - 二酰基甘油合成CL的最后一步。CL及其合成主要定位于线粒体内膜，并且通常认为CL是许多线粒体过程的必需成分。通过对可能编码磷脂生物合成酶的基因进行同源性搜索，我们克隆了酿酒酵母中编码CL合酶的基因（CLS1）。在其内源启动子或酵母中可诱导的GAL1启动子下过表达CLS1基因，以及在杆状病毒感染的昆虫细胞中表达CLS1，均导致CL合酶活性升高。在单倍体酵母菌株中破坏CLS1基因导致CL合酶活性丧失、无法检测到CL、磷脂酰甘油水平升高5倍，以及对CL具有高亲和力的染料无法对线粒体进行染色。cls1::TRP1缺失突变体在可发酵和不可发酵碳源上均能生长，但在后者上生长较差。细胞色素c氧化酶的水平和活性正常，并且一种其积累依赖于膜质子电化学势的染料能够有效地对线粒体进行染色。这些结果明确鉴定了酵母中编码CL合酶的基因。
