Dong Xinyan, Liu Manni, Liu Xue, Liu Min, Zhang Xiangling, Wang Guoqing
West China School of Public Health and West China Fourth Hospital, Sichuan University, Chengdu 610041, China.
Wei Sheng Yan Jiu. 2020 Sep;49(5):785-822. doi: 10.19813/j.cnki.weishengyanjiu.2020.05.016.
Establish the prokaryotic expression system of Amuc_1100 protein from Akkermansia muciniphila, and analyze the effects of this protein on the body weight, blood glucose, intestinal barrier function and Akkermansia muciniphila abundance in rats fed with high-fat diet combined streptozotocin(STZ)injection.
PCR product of Amuc_1100 Gene(Gene ID: 34174504) was linked to the double enzyme-digested pET-26 b plasmid vector. The recombinant expression plasmid pET-26 b-Amuc_1100 then transformed into E. Coli BL21. The verified clones through sequence analysis were induced by the addition of IPTG. High-fat diet rats were interfered with the purified protein at high and low doses. The changes of body weight, blood glucose, intestinal barrier function and Akkermansia muciniphila abundance were analyzed.
The recombinant expression plasmid pET-26 b-Amuc_1100 was successfully constructed. The sequencing result showed 100% similarity to the Amuc_1100 gene in GenBank. The molecular weight of the protein obtained was 42 kDa. The intervention of Amuc_1100 protein can reduce the weight of rats fed with high-fat diet combined STZ injection to some extent, improve barrier function and increase the abundance of Akkermansia muciniphila in intestine.
The prokaryotic expression system of Amuc_1100 protein was successfully constructed, which has a certain regulatory effect on rats fed with high-fat diet combined STZ injection.
建立嗜黏蛋白阿克曼氏菌Amuc_1100蛋白的原核表达系统，并分析该蛋白对高脂饮食联合链脲佐菌素（STZ）注射的大鼠体重、血糖、肠道屏障功能及嗜黏蛋白阿克曼氏菌丰度的影响。
将Amuc_1100基因（基因ID：34174504）的PCR产物与经双酶切的pET-26 b质粒载体连接。将重组表达质粒pET-26 b-Amuc_1100转化至大肠杆菌BL21中。经序列分析验证的克隆通过添加异丙基-β-D-硫代半乳糖苷（IPTG）进行诱导。高脂饮食大鼠分别用高、低剂量的纯化蛋白进行干预。分析体重、血糖、肠道屏障功能及嗜黏蛋白阿克曼氏菌丰度的变化。
成功构建了重组表达质粒pET-26 b-Amuc_1100。测序结果显示与GenBank中的Amuc_1100基因相似度为100%。获得的蛋白分子量为42 kDa。Amuc_1100蛋白干预可在一定程度上降低高脂饮食联合STZ注射大鼠的体重，改善屏障功能，并增加肠道中嗜黏蛋白阿克曼氏菌的丰度。
成功构建了Amuc_1100蛋白的原核表达系统，该系统对高脂饮食联合STZ注射的大鼠具有一定的调节作用。
