2-酮基葡萄糖酸（2KGA）是食品添加剂D-异抗坏血酸（钠）生产的关键中间体，国内外工业生产均采用发酵法。在发酵过程中，2KGA既是发酵的目的产物，也是可被生产菌株利用的中间代谢产物。因此，如何有效减少发酵液中的2KGA被转运至胞内成为进一步提升2KGA发酵水平的重要课题。本项目拟对工业生产菌株变形假单胞菌JUIM01的2KGA转运蛋白KguT进行异源表达，并对其结构特征、转运特性及其编码基因在发酵过程中转录水平上的变化进行系统分析；然后，通过研究2KGA代谢的调控因子PtxS与靶DNA及效应物之间的相互作用，解析KguT的基因转录调控机制；最终，基于研究得到的KguT的转运特性及其基因转录调控机制，采用定点突变或基因敲除技术对KguT的编码基因、启动子区等进行改造，结合考察菌株生理代谢的变化，实现对菌体细胞2KGA转运过程的理性调控，得到2KGA分解代谢显著弱化的2KGA高产菌株。

2-Keto-gluconic acid (2KGA) is the key intermediate for the manufacture of a safe food additive erythorbic acid (isoascorbic acid), and industrially produced by microbial wordwidely. Our previous study revealed that 2KGA is the target product and also the intermediate metabolite during the fermentation process. Hence how to efficiently decrease the intracellular transportation of 2KGA in the broth is the key task for improving the 2KGA fermentation yield and industrial benefits. The present proposal will focus on: 1) heterologously expressing the 2KGA transporter (KguT) of the industrail producer Pseudomonas plecoglossicida JUIM01, elucidating its structural characteristics and 2KGA-transporting models, and analyzing changes of KguT gene at transcription level; 2) elucidating the transcriptional regulation mechanism of KguT gene by investigating the intereaction of regulator PtxS, target DNA and effectors, and 3) developing and selecting the mutants with significantly lowered transportation of 2KGA by reconstruction of KguT gene or promoter regions using site-directed mutagenesis or gene knock-out methods, and comparison of the cell growth and 2KGA production/yield/producivity based on the 2KGA-transporting characteristics and regulation mechanism.