Phenol Production By Metabolically Engineered Escherichia coli

Engineering Escherichia coli for biological production of phenol is considered infeasible due to its toxicity and complex biosynthetic network. To address these challenges, we took advantage of the synthetic sRNA-based gene knockdown technology to simultaneously engineer 18 E. coli strains and screen them for higher phenol tolerance and precursor tyrosine availability. The knock-down of csrA and tyrR genes were identified to be crucial for redirecting carbon flux toward tyrosine and applied to all the E. coli strains with over-expression of the key genes for the biosynthesis of tyrosine and phenol (tyrosine phenol-lyase). The engineered strains showed substantially different metabolic phenotypes. The engineered BL21 (DE3) strain produced 419 mg/L of phenol in flask culture and 1.69 g/L in fed-batch fermentation. A biphasic fed-batch fermentation using glycerol tributyrate was develope for in situ extraction while minimizing the phenol toxicity. The preferential partition of phenol from culture medium into glycerol tributyrate increased the titer and productivity to 3.79 g/L and 0.18 g/L/h, respectively. [This work was supported by the Intelligent Synthetic Biology Center through the Global Frontier Project (2011-0031963) of the Ministry of Science, ICT & Future Planning through the National Research Foundation of Korea.]