作　　者： ; ; ; ; ; ; ; ;

机构地区： 华南农业大学兽医学院

出　　处： 《畜牧与兽医》 2013年第9期1-5,共5页

摘　　要： 酰基甘油酯酶(MAGL)是将酰基甘油分解为甘油和游离脂肪酸的丝氨酸水解酶家族成员之一,在酯代谢中起着关键酶的作用,是研制抗鸡球虫药物的重要靶标。本研究利用生物信息学技术预测拼接了柔嫩艾美耳球虫MAGL基因序列,以第二代裂殖子总RNA为模板,通过RTPCR技术获得Etmagl基因。将Etmagl与pCold-43a载体连接,构建pCold-43a-Etmagl重组载体,并在大肠杆菌BL21中获得可溶性蛋白,经亲和层析获得纯化的重组蛋白。结果显示,扩增的Etmagl序列ORF长1 752 bp,编码584个氨基酸,与预测序列相似度为99%,与弓形虫MAGL相似度好(55%);IPTG诱导后融合蛋白高效表达,大小约为114 ku,经免疫印迹鉴定为目的蛋白。本研究成功利用大肠埃希菌原核表达体系重组表达并纯化了柔嫩艾美耳球虫酰基甘油脂肪酶,为建立以MAGL为靶标的抗球虫药物筛选模型奠定了基础。 Monoacylglycerol lipase （MAGL）, a member of serine hydrolase family as the key enzyme in ester metabolism that hydrolyzes acylglyeerols to glycerol and fatty acids, could be used as novel drug target for anti-coccidial drugs. The MAGL sequence of Eimeria tenella was predicted by bioinformatics and electronic cloning technology, and the gene fragment was amplified by RT-PCR using total RNA from the second schizonts of E. tenella as template. The Etmagl gene was ligated with pCold-43a vector, and then transformed into E. coli BL21 （ DE3 ） to express. Soluble protein was purified by the affinity chromatography assay. The result showed that the Etmagl ORF is 1 752 bp in length, coding 584 aa peptide. The sequence similarity is 99% to prediction, but only 55% to that of Toxoplasma gondii. The fusion protein with molecular weight of 114 ku was efficiently expressed in BL21 （ DE3 ） with the induction of IPTG. This study lays a basis for building a targeted EtMAGL model to screen a new anti-coccidial drug.

关 键 词： 酰基甘油酯酶 柔嫩艾美耳球虫 表达 纯化

领　　域： [农业科学] [农业科学] [农业科学]