Oishi H, Tsuda S, Watanabe Y, Tamai Y
Department of bioresources, Faculty of Agriculture, Ehime University, Japan.
Biosci Biotechnol Biochem. 1996 Jul;60(7):1087-92. doi: 10.1271/bbb.60.1087.
Phospholipase B from Schizosaccharomyces pombe was purified by ammonium sulfate fractionation and chromatographed on phenyl-Sepharose CL-4B, DEAE-Toyopearl 650M, and TSK gel G4000SW columns. The purified enzyme was a glycoprotein with molecular weight of approximately 300,000 and 100,000-150,000 by gel filtration and SDS-polyacrylamide gel electrophoresis, respectively. The isoelectric point was pH 4.7. The optimum pH of the enzyme was 2.5 and no activity was detected at neutral and alkaline pHs. The enzyme was not heat-stable. Enzyme activity was slightly stimulated by divalent ions except Fe2+ and 0.1% sodium deoxycholate, and inhibited by Fe2+, Fe3+, 0.1% sodium dodecyl sulfate, and 0.01% cetyltrimethylammonium bromide. The enzyme hydrolyzed mono- and diacylphospholipids, and phosphatidylinositol was hydrolyzed most preferentially. Triglyceride was not hydrolyzed. The enzyme also had acyltransferase activity on lysophosphatidylcholine, forming the corresponding diacylphosphatidylcholine.
通过硫酸铵分级分离法对来自粟酒裂殖酵母的磷脂酶B进行纯化，并在苯基琼脂糖凝胶CL - 4B、DEAE - 琼脂糖凝胶650M和TSK凝胶G4000SW柱上进行色谱分析。通过凝胶过滤和SDS - 聚丙烯酰胺凝胶电泳测定，纯化后的酶是一种糖蛋白，分子量分别约为300,000和100,000 - 150,000。其等电点为pH 4.7。该酶的最适pH为2.5，在中性和碱性pH条件下未检测到活性。该酶不耐热。除Fe2 +和0.1%脱氧胆酸钠外，二价离子对酶活性有轻微刺激作用，而Fe2 +、Fe3 +、0.1%十二烷基硫酸钠和0.01%十六烷基三甲基溴化铵对酶活性有抑制作用。该酶可水解单酰基和二酰基磷脂，其中磷脂酰肌醇最易被水解。甘油三酯不被水解。该酶对溶血磷脂酰胆碱还具有酰基转移酶活性，可形成相应的二酰基磷脂酰胆碱。
