Enhanced tyrosine production in Corynebacterium glutamicum via AroG A178V, RNase J deficiency, and carbon flux...

Tyrosine production in Corynebacterium glutamicum has mainly been improved because of the use of feedback-resistant enzymes and redirection of carbon flux. However, the genetic basis of the classical feedback-deregulated phenotype and contribution of post-transcriptional regulation remain unclear. In this study, we identified the mutation responsible for the feedback-deregulated phenotype of the aroG-csm region in the phenylalanine-producing C. glutamicum strain KY10694, examined the effect of RNase J deficiency on tyrosine production, and developed a stepwise strategy to further improve tyrosine production. Sequence analysis revealed a single nucleotide substitution in aroG that resulted in an A178V amino acid substitution, whereas no mutation was detected in csm. Enzyme assays showed that the KY10694-derived aroG-csm gene products converted the feedback response of chorismate mutase activity to phenylalanine and tyrosine. In a 2 L jar fermentor cultivation, the strain harboring the KY10694-derived aroG-csm genes produced 5.45 mM tyrosine, and deletion of rnj, which encodes RNase J, increased tyrosine production to 10.0 mM. Transcriptome analysis of the RNase J-deficient strain revealed broad changes in the expression of genes involved in the central carbon metabolism and aromatic amino acid biosynthesis. Further redirection of carbon flux by deletion of pheA and subsequent deletion of ppc, which encode prephenate dehydratase and phosphoenolpyruvate carboxylase, respectively, increased tyrosine production to 21.7 and 24.8 mM, respectively. These results demonstrate that integrating pathway deregulation through AroG A178V, regulatory rewiring through RNase J deficiency, and rational carbon flux redirection is an effective strategy for improving tyrosine production in C. glutamicum.