重组甜味蛋白Monellin在E.coli中的表达条件及其对ICR小鼠血糖的影响  

Expression Conditions of Recombinant Sweet Protein Monellin in E.coli and Its Effect on Blood Glucose in ICR Mice

作　　者：杨冬梅 王晨璐 刘嘉健 方越 葛瑛 韩淳致 曹荣月[1] YANG Dong-mei;WANG Chen-lu;LIU Jia-jian;FANG Yue;GE Ying;HAN Chun-zhi;CAO Rong-yue(School of Life Science and Technology,China Pharmaceutical University,Nanjing 211198,China;International Academy of Chinese Medicine,University of Macao,Macao 00853,China)

机构地区：[1]中国药科大学生命科学与技术学院,南京211198 [2]澳门大学国际中华医药研究院,中国澳门00853

出　　处：《中国调味品》2021年第7期1-6,共6页China Condiment

基　　金：2020互联网大创“牌过留痕,生化不再危机”(3312000657);2019年校级创新创业课程;国家级大学生创新创业训练计划项目(J1310032,202010316014G);国家自然科学基金资助项目(81673340);中国药科大学2020年创新创业菁英培育计划立项“35糖尿病患者的甜”。

摘　　要：实验探究了重组Monellin菌株的最佳培养条件,并对其进行了甜味阈值确认和血糖值影响实验。实验所用菌种为实验室构建保存的BL21(DE3)-pET28a Monellin菌种,利用不同浓度的乳糖和IPTG诱导该菌种生成目的蛋白,探究其最佳诱导条件,利用镍柱亲和层析纯化目的蛋白后进行相对甜度和甜味阈值实验,随后设置甜度梯度,对小鼠进行灌胃比较蔗糖和Monellin的升血糖作用及不同浓度Monellin对血糖的影响。实验结果表明,0.9 mmol/L IPTG诱导5 h的Monellin表达量最多,其甜味阈值为100μg/mL,相对甜度为蔗糖的800倍。灌胃结果表明,Monellin升血糖作用不显著,并且在一定范围内对血糖的影响不随其浓度的改变而发生变化。该实验为Monellin在糖尿病等代糖食品市场上的应用提供了科学的理论依据。The optimal culture conditions of recombinant Monellin strain are investigated,and the sweetness threshold is confirmed and the effect of blood glucose is tested.The strain used in the experiment is BL21(DE3)-pET28a Monellin strain constructed and preserved in the laboratory.Different content of lactose and IPTG is used to induce the strain to produce the target protein,and the optimal induction conditions are investigated.After purifying the target protein by nickel column affinity chromatography,the relative sweetness degree and sweetness threshold are tested,and then the sweetness gradient is set.The mice are given intragastric gavage to compare the glycemic effects of sucrose and Monellin and the effect of different content of Monellin on blood glucose.The results show that Monellin expression amount induced by 0.9 mmol/L IPTG for 5 h is the highest,and its sweetness threshold is 100μg/mL,and its relative sweetness degree is 800 times that of sucrose.The results of intragastric gavage show that Monellin has no significant effect on blood glucose,and its effect on blood glucose does not change with the change of its concentration in a certain range.This experiment has provided a scientific theoretical basis for the application of Monellin in the market of sugar substitutes such as diabetes.

关 键 词：甜味蛋白Monellin 诱导条件 镍柱亲和层析 甜味阈值 血糖值 

分 类 号：TS201.21[轻工技术与工程—食品科学]