基于紫色杆菌素生物合成途径的L-色氨酸生物传感器的构建  

Development of an L-tryptophan Biosensor Based on the Violacein Biosynthesis Pathway

作　　者：李仁瀚 张乐乐[1,2] 刘春立 刘秀霞 白仲虎[1,2,3] 杨艳坤 李业 LI Ren-han;ZHANG Le-le;LIU Chun-li;LIU Xiu-xia;BAI Zhong-hu;YANG Yan-kun;LI Ye(The Key Laboratory of Industrial Biotechnology,Ministry of Education,Jiangnan University,Wuxi 214122;National Engineering Research Center for Cereal Fermentation and Food Biomanufacturing,Jiangnan University,Wuxi 214122;Jiangsu Provincial Engineering Research Center for Bioactive Product Processing,Jiangnan University,Wuxi 214122)

机构地区：[1]江南大学工业生物技术教育部重点实验室,无锡214122 [2]江南大学粮食发酵与食品生物制造国家工程研究中心,无锡214122 [3]江苏省生物活性制品加工工程技术研究中心,无锡214122

出　　处：《生物技术通报》2023年第10期80-92,共13页Biotechnology Bulletin

基　　金：国家自然科学基金面上项目(21878124)。

摘　　要：结合生物传感器进行高通量筛选是鉴定L-色氨酸高产菌株的有力工具,但现有的L-色氨酸生物传感器普遍存在工作范围和动态范围都较低的缺点。在大肠杆菌中表达紫色杆菌素生物合成途径,测试不同来源的VioA酶,结合RBS工程,开发了一种新型酶偶联L-色氨酸生物传感器。测试发现来自Chromobacterium violaceum的VioA酶可使该传感器的检测上限扩大至10 g/L外源添加的L-色氨酸;将其RBS的初始翻译速率降低到约2000时,传感器的动态范围扩大到55倍;通过肉眼可直观地区分L-色氨酸产量不同的大肠杆菌菌株。这种新型的L-色氨酸生物传感器可结合高通量筛选等手段,在鉴定L-色氨酸及其高附加值衍生物高产菌株等方面发挥巨大作用。High-throughput screening combining with biosensors is a powerful tool for identifying high-yield L-tryptophan producing strains,but current L-tryptophan biosensors generally have the disadvantage of low operational and dynamic ranges.By expressing the violacein biosynthetic pathway in Escherichia coli and testing different sources of Vio A enzyme and RBS engineering,a novel enzyme-coupled L-tryptophan biosensor was developed.It was found that the VioA enzyme from Chromobacterium violaceum expanded detection limit of the biosensor up to 0-10 g/L of exogenously added L-tryptophan.Reducing its translation initiation rate to approximately 2 000 expanded the dynamic range of the biosensor by 55-fold.Different strains of E.coli with varying L-tryptophan yields could be visually distinguished with the naked eye.This novel L-tryptophan biosensor can play a significant role in identifying high-yield L-tryptophan and its high-value derivatives producing strains through combining methods such as high-throughput screening.

关 键 词：大肠杆菌 紫色杆菌素 L-色氨酸 生物传感器 RBS工程 

分 类 号：R313[医药卫生—基础医学] TP212.3[自动化与计算机技术—检测技术与自动化装置]