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

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.