磷脂酰胆碱特异性和磷脂酰肌醇特异性磷脂酶C酶学性质及其在油脂脱胶中的应用

摘要

通过基因工程方法将具有磷脂酰胆碱 (PC) 、磷脂酰乙醇胺 (PE) 特异性磷脂酶C (PC-PLC) 和磷脂酰肌醇 (PI) 特异性磷脂酶C (PI-PLC) 基因分别在枯草芽孢杆菌WB800中进行胞外表达.对2种重组蛋白粗酶的酶学性质进行研究, 并探究其用于油脂脱胶中的效果.结果表明:PC-PLC和PI-PLC均在30～45℃和中性偏酸的条件下具有较好的稳定性, 2 h后残余活性仍在60％以上.Zn2+、Mg2+对PC-PLC的活性有促进作用, 而Ca2+对PI-PLC的活性有促进作用.油脂脱胶研究结果表明, PC-PLC的最适脱胶条件为50℃、pH 6.0、反应2 h、酶添加量为80 mg/kg;PI-PLC的最适脱胶条件为45℃、pH 7.0、反应2 h、酶添加量为60 mg/kg.PC-PLC和PI-PLC联合脱胶实验中大豆毛油中磷含量从531 mg/kg降至4.1 mg/kg, 此时甘油二酯 (DAG) 含量增加0.95％.%We transported two phospholipase C genes to Bacillus sublitis WB800 to express phosphatidylcholine, phosphatidylethanolamine-and phosphatidylinositol-specific properties by genetic engineering methods.Then the two recombinant proteins were identified, characterized and evaluated on their degumming efficiency.Both enzymes exhibited high temperature (retaining 60％ catalytic activity, 30-45 ℃ for 2 h) and pH stability (retaining 60％ activity, pH 6.0-7.0 for 2 h).Besides, the metal ions (Zn2+ and Mg2+) could increase the catalytic activity of PC-PLC whereas the activity of PI-PLC increased by Ca2+.The oil degumming results showed that the optimal degumming condition of PC-PLC was 50 ℃, pH 6.0, enzyme dosage of 80 mg/kg oil and reaction time of 2.5 h.The optimal degumming condition of PI-PLC was 45 ℃, pH 7.0, enzyme dosage of 60 mg/kg oil, reaction time of 2.5 h.The degumming effects of combining PC-PLC and PI-PLC to treat crude soybean oil, phosphorus content decreased from 531 mg/kg to 4.1 mg/kg and the yield of DAG increased 0.95％.