植物乳杆菌肌球交叉反应抗原MCRA的克隆表达及功能鉴定

Cloning, expression and enzymatic activity of the myosin cross reactive antigen gene from Lactobacillus plantarum ZS2058

摘要: 根据NCBI中已报道亚油酸异构酶基因的序列特征从植物乳杆菌ZS2058克隆获得了产CLA的关键酶基因肌球交叉反应抗原MCRA,构建表达载体后,在大肠杆菌中实现了表达,SDS-PAGE和Western Blot检测结果为胞内可溶表达,其大小约为67ku,将MCRA蛋白进行亲和层析后进行功能鉴定,GC-MS结果显示此蛋白能将亚油酸转化为羟基化衍生物10羟基-顺12-十八碳烯酸,将油酸转化为10-羟基-十八碳酸。

Abstract: Multiple enzymes were involved for CLA production in lactic acid bacteria (LAB) . Myosin cross reactive antigen (MCRA) gene was amplified with the specific primer from Lactobacillus plantarum ZS2058 according to reported putative linoleate isomerease in LAB in NCBI. MCRA gene was cloned and expressed successfully into Escherichia coli, the recombinant MCRA was analyzed by SDS-PAGE and Western Blot, and the results indicated the molecular weight of MCRA was 67ku. After purification with affinity chromatography, the activity of recombinant MCRA was analyzed. The GC-MS results showed that MCRA catalyzed linoleic acid into 10-hydroxy-cis-12-octadecenoic acid, and catalyzed oleic acid into 10-hydroxy-octadecanoic acid.