Sanjanwala M, Sun G Y, MacQuarrie R A
Division of Molecular Biology and Biochemistry, School of Basic Life Sciences, University of Missouri-Kansas City 64110.
Biochim Biophys Acta. 1989 Nov 28;1006(2):203-8. doi: 10.1016/0005-2760(89)90197-5.
Heart muscle microsomes catalyze the transacylation of lysophosphatidylcholine (lyso PC) to produce phosphatidylcholine (PC). The enzyme which catalyzes this reaction, lyso PC:lyso PC transacylase, has been isolated and characterized from bovine heart muscle microsomes. The purification of the enzyme was achieved by a procedure involving extraction with 3-[3-cholamidopropyl)dimethylammonio)-1-propanesulfonate (CHAPS) detergent and chromatography on DEAE-cellulose, Reactive blue agarose, and Matrex gel green A. The purified enzyme was nearly homogeneous and consisted of a single molecular species of 128 kDa as determined by polyacrylamide gel electrophoresis in the presence of dodecyl sulfate. The catalytic activity of the enzyme was dependent on the presence of either CoA or acyl-CoA, both of which maximally stimulated at concentrations of approx. 10 microM. Analysis of the PC produced in the reaction showed that the enzyme catalyzed a transacylation in which both acyl groups arose from lyso PC. Furthermore, the enzyme did not possess acyl-CoA:lyso PC acyltransferase activity, lysophospholipase or acyl-CoA hydrolase activity, nor did it catalyze transacylation from lyso PC to lysophosphatidylethanolamine, lysophosphatidylinositol or lysophosphatidylserine. Although transacylation was highly specific for lyso PC as the substrate, various unsaturated fatty acyl-CoA derivatives served as activators. Palmitoyl-CoA and stearoyl-CoA did not significantly activate, although acetyl-CoA was an effective activator. Further modulation of activity was produced by palmitic acid and PC, both of which further activated the enzyme in the presence of oleoyl-CoA, whereas arachidonic acid, oleic acid, phosphatidylethanolamine and phosphatidylserine had no effect on activity. The high activity of this transacylase and its regulation by lipids suggests an important role for disaturated PC species in membranes and a mechanism for controlling the metabolism of lyso PC.
心肌微粒体催化溶血磷脂酰胆碱（lyso PC）的转酰基作用以产生磷脂酰胆碱（PC）。催化此反应的酶，即lyso PC:lyso PC转酰基酶，已从牛心肌微粒体中分离并进行了特性鉴定。该酶的纯化通过以下步骤实现：用3-[(3-胆酰胺丙基)二甲基铵基]-1-丙烷磺酸盐（CHAPS）去污剂提取，然后在DEAE-纤维素、活性蓝琼脂糖和Matrex凝胶绿A上进行色谱分离。纯化后的酶几乎是均一的，通过十二烷基硫酸钠存在下的聚丙烯酰胺凝胶电泳测定，其由一种分子量为128 kDa的单一分子物种组成。该酶的催化活性依赖于辅酶A（CoA）或酰基辅酶A（acyl-CoA）的存在，二者在约10 microM的浓度下均能产生最大刺激作用。对反应中产生的PC进行分析表明，该酶催化的转酰基作用中，两个酰基均来自lyso PC。此外，该酶不具有酰基辅酶A:lyso PC酰基转移酶活性、溶血磷脂酶或酰基辅酶A水解酶活性，也不催化从lyso PC到溶血磷脂酰乙醇胺、溶血磷脂酰肌醇或溶血磷脂酰丝氨酸的转酰基作用。尽管转酰基作用对lyso PC作为底物具有高度特异性，但各种不饱和脂肪酰基辅酶A衍生物可作为激活剂。棕榈酰辅酶A和硬脂酰辅酶A没有显著激活作用，尽管乙酰辅酶A是一种有效的激活剂。棕榈酸和PC对活性产生进一步调节作用，二者在油酰辅酶A存在下均能进一步激活该酶，而花生四烯酸、油酸、磷脂酰乙醇胺和磷脂酰丝氨酸对活性没有影响。这种转酰基酶的高活性及其受脂质调节的特性表明，二饱和PC物种在膜中具有重要作用，并且存在一种控制lyso PC代谢的机制。
