Whole-cell Catalyzed Production of L-tyrosine in Recombinant Bacillus subtilis

Objective: This study conducted a recombinant Bacillus subtilis strain capable of producing L-Tyr utilizing phenol, sodium pyruvate, and ammonia. Moreover, the conditions of cell culture and catalytic reaction were optimized to improve L-Tyr production. Methods: The tyrosine phenol-lyase (TPL) gene from Pantoea agglomerans was codon-optimized and successfully over-expressed in B. subtilis. The conditions of induction and whole-cell transformation were optimized using single-factor experiments for L-Tyr production. Results: The highest TPL enzyme activity was achieved 4.65±0.15 U·g⁻¹ in the recombinant B. subtilis at 20 ℃ with 2.0 g/L xylose after 36 hours. Under the whole-cell transformation conditions of 75 mmol/L phenol, 75 mmol/L sodium pyruvate, 487 mmol/L ammonium chloride, 2.0 g/L sodium sulfite, 2.0 g/L EDTA, 0.08 g/L pyridoxal phosphate (PLP), wet cell mass of 50 g/L, pH8, and 35 ℃, the L-Tyr production increased to 9.38 g/L with a conversion rate of 73.24%. Coping with the issue of decreased TPL enzyme activity caused by phenol resistance, a batch feeding strategy was implemented during the whole-cell transformation, resulting in a final L-Tyr production of 15.12 g/L with a conversion rate of 75.51% after 20 hours. Conclusion: These findings demonstrate the successful conversion of phenol and sodium pyruvate into L-Tyr by the recombinant B. subtilis strain. The study would provide a theoretical and technical foundation for the production of food-grade L-Tyr using whole-cell biocatalysis, highlighting its potential for practical applications.