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l-DOPA is a major drug for the treatment of Parkinson’s disease, and microbial enzymatic synthesis is an essential way for its industrial production. The expression of tyrosine phenol-lyase (TPL) derived from Erwinia herbicolaunder six different constitutive promoters in Escherichia coliwas studied. The recombinant strain 5 can express TPL well with growth. The optimal medium composition of this strain was as follows: tryptone 18 g/L, yeast extract 28 g/L, glycerol 10 g/L. After the cell culture is completed, the collected cells were used to catalyze the production of l-DOPA, and the concentration can reach 67.9 g/L.