Kelly B L, Greenberg M L
Cellular and Molecular Biology Program, University of Michigan, Ann Arbor 48109-0606.
Gene. 1994 Sep 15;147(1):111-4. doi: 10.1016/0378-1119(94)90048-5.
Cardiolipin (CL) is a structurally unique phospholipid having important functional roles in both prokaryotic and eukaryotic cells. The genes encoding CL biosynthetic enzymes have been identified and extensively studied in Escherichia coli, and manipulation of CL biosynthesis in this organism has elucidated a great deal about CL function in prokaryotes. In contrast, little is known about CL biosynthesis or its regulation in eukaryotic cells. We sought to determine whether we could utilize E. coli genes to manipulate expression of CL biosynthetic enzymes and CL content in yeast. The E. coli pgsA gene encodes phosphatidylglycerophosphate synthase (PGPS), catalyzing the first step in the CL biosynthetic pathway. We constructed plasmids with pgsA under the control of the yeast CUP1 promoter. Extracts of Saccharomyces cerevisiae cells transformed with this plasmid contained high levels of E. coli PGPS activity. However, when compared to cells transformed with a control plasmid, pgsA-transformed cells did not exhibit differences in phospholipid composition. The most likely explanation is that the in vitro activity of the E. coli pgsA product is not indicative of its activity in vivo, due to mislocalization of the enzyme and/or inaccessibility of the enzyme to the substrates. To our knowledge, this is the first demonstration of expression of a bacterial phospholipid biosynthetic enzyme in yeast.
心磷脂（CL）是一种结构独特的磷脂，在原核细胞和真核细胞中都具有重要的功能作用。编码CL生物合成酶的基因已在大肠杆菌中被鉴定并广泛研究，对该生物体中CL生物合成的操纵已阐明了许多关于CL在原核生物中的功能。相比之下，关于CL在真核细胞中的生物合成或其调控知之甚少。我们试图确定是否可以利用大肠杆菌基因来操纵酵母中CL生物合成酶的表达和CL含量。大肠杆菌pgsA基因编码磷脂酰甘油磷酸合酶（PGPS），催化CL生物合成途径的第一步。我们构建了在酵母CUP1启动子控制下带有pgsA的质粒。用该质粒转化的酿酒酵母细胞提取物含有高水平的大肠杆菌PGPS活性。然而，与用对照质粒转化的细胞相比，pgsA转化的细胞在磷脂组成上没有表现出差异。最可能的解释是，由于酶的错误定位和/或酶对底物的不可及性，大肠杆菌pgsA产物的体外活性不能指示其体内活性。据我们所知，这是首次在酵母中表达细菌磷脂生物合成酶的证明。
