Tamai K T, Greenberg M L
Department of Biological Chemistry, University of Michigan, Ann Arbor 48709-0606.
Biochim Biophys Acta. 1990 Sep 18;1046(2):214-22. doi: 10.1016/0005-2760(90)90192-z.
Cardiolipin (CL) synthase activity was characterized in mitochondrial extracts of the yeast Saccharomyces cerevisiae and was shown for the first time to utilize CDP-diacylglycerol as a substrate. CL synthase exhibited a pH optimum of 9.0. Maximal activity was obtained in the presence of 20 mM magnesium with a Triton X-100: phospholipid ratio of 1:1. The apparent Km values for phosphatidylglycerol and CDP-diacylglycerol were 1 mM and 36 microM, respectively. CL synthase activity was maximal at 45 degrees C and heat inactivation studies showed that the enzyme retained greater than 75% of its activity at temperatures up to 55 degrees C. To study the regulation of CL synthase, the enzyme was assayed in cells grown under conditions known to affect general phospholipid synthesis. Unlike many phospholipid biosynthetic enzymes including PGP synthase, which catalyzes the initial step in CL biosynthesis, CL synthase was not repressed in cells grown in the presence of the phospholipid precursor inositol. Detailed procedures for the enzymatic synthesis of 32P-labelled substrates are described.
在酿酒酵母的线粒体提取物中对心磷脂（CL）合酶活性进行了表征，首次证明其以CDP - 二酰甘油为底物。CL合酶的最适pH为9.0。在20 mM镁存在下，Triton X - 100与磷脂比例为1:1时获得最大活性。磷脂酰甘油和CDP - 二酰甘油的表观Km值分别为1 mM和36 μM。CL合酶活性在45℃时最大，热失活研究表明该酶在高达55℃的温度下仍保留其活性的75%以上。为了研究CL合酶的调控，在已知影响一般磷脂合成的条件下生长的细胞中对该酶进行了测定。与许多磷脂生物合成酶不同，包括催化CL生物合成第一步的PGP合酶，CL合酶在磷脂前体肌醇存在下生长的细胞中不会被抑制。描述了32P标记底物酶促合成的详细步骤。
