Optimal Strain and Knockdown Target Screening Method Using Synthetic Small Regulatory RNA for Enhanced Production of Tyrosine and Cadaverine

Abstract

Small regulatory RNAs (sRNAs) are gene expression regulators which acton post-transcriptional phase of bacterial gene expression. Here, we developedsynthetic small regulatory RNA system to down-regulate the gene expression.The target mRNA binding region of MicC, one of the well-studied sRNAs inEscherichia coli, was replaced to translation initiation sequence of our targetgenes. We found that MicC scaffold based synthetic sRNA is properly repressedexpression of DsRed2, a fluorescence protein. Synthetic sRNAs was used formetabolic engineering to enhance the production of tyrosine and cadaverine.Through screening of 14 different strains which harboring one or combinationof the synthetic sRNAs targeting ppc, tyrR, csrA, and pgi, we isolated a tyrosineproducer producing 2 g per liter of tyrosine. Using a library of 130 syntheticsRNAs, we screened knockdown targets that increase cadaverine productivitysubstantially. Repression of murE led to a 55% increase in cadaverine productioncompared to the base strain. [This work was supported by the TechnologyDevelopment Program to Solve Climate Changes on Systems MetabolicEngineering for Biorefineries; theIntelligent Synthetic Biology Center through the Global Frontier Project of the Ministry of Science, ICT and Future Planning (MSIP)through the National Research Foundation of Korea]