重组磷脂酶D Sp-TLI146的发掘及其在DHA磷脂酰单甘油酯的合成中的应用  

Discovery of Recombinant Phospholipase D Sp-TLI146 and Its Application in Synthesis of DHA Phosphatidyl Monoglycerides

作　　者：贺晨曦 陈曦 刁汝静 孙建安[1,2,3] 毛相朝 HE Chenxi;CHEN Xi;DIAO Rujing;SUN Jian'an;MAO Xiangzhao(College of Food Science and Engineering,Ocean University of China,Qingdao 266003,Shandong,China;Qingdao Key Laboratory of Food Biotechnology,Qingdao 266404,Shandong,China;Key Laboratory of Biological Processing of Aquatic Products,China National Light Industry,Qingdao 266404,Shandong,China)

机构地区：[1]中国海洋大学食品科学与工程学院,山东青岛266003 [2]青岛市食品生物技术重点实验室,山东青岛266404 [3]中国轻工业水产品生物加工重点实验室,山东青岛266404

出　　处：《食品研究与开发》2024年第21期187-194,216,共9页Food Research and Development

基　　金：国家自然科学基金项目(32172165);国家重点研发计划课题(2019YFD0901901)。

摘　　要：从链霉菌中克隆一个磷脂酶D(phospholipase D,PLD)的编码基因Sp-TLI146,并将其在大肠杆菌BL21(DE3)中进行异源表达。利用镍离子亲和层析后得到纯酶并对其酶学性质进行研究。酶学性质研究显示,重组PLD的最适温度为65℃,在50~65℃时,其相对酶活力仍高于60%;最适pH值为6.0的磷酸-磷酸钠缓冲液,在pH4.0~8.0的相对酶活力高于70%。以鱿鱼卵为来源,使用有机溶剂浸提法提取DHA磷脂酰胆碱(docosahexaenoic acid phosphatidylcholine,DHAPC)并对其进行纯化,在重组PLD Sp-TLI146的催化作用下,与不同的甘油酯反应合成DHA磷脂酰单甘油酯,在最适条件下,反应4 h转化率可达90%以上。A gene encoding of phospholipase D(PLD),Sp-TLI146,was cloned from Streptomyces sp.and heterologously expressed in Escherichia coli BL21(DE3).After affinity chromatography with nickel ion,the pure enzyme was obtained and its enzymatic properties were investigated.According to enzymatic property studies,the optimum temperature of recombinant PLD was 65℃,while the relative enzyme activity remained higher than 60% at 50-65℃.The optimum pH value was phosphate-sodium phosphate buffer at pH6.0,while the relative enzyme activity reached higher than 70% at pH 4.0-8.0.DHA phosphatidylcholine(docosahexaenoic acid phosphatidylcholine,DHAPC)was extracted from squid eggs with organic solvent extraction and purified.DHA phosphatidyl monoglycerides were synthesized by reacting with different glycerides under the catalytic effect of recombinant PLD Sp-TLI146.The conversion rate of the reaction for 4 h could reach more than 90% under the optimal conditions.

关 键 词：磷脂酶D 异源表达 酶学性质 DHA磷脂酰单甘油酯 转酯反应 

分 类 号：TS201.25[轻工技术与工程—食品科学] TS221[轻工技术与工程—食品科学与工程]