A bi-enzymatic cascade to yield pyruvate as co-substrate for l -tyrosine production
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BIOTECHNOLOGICAL PRODUCTS AND PROCESS ENGINEERING
A bi-enzymatic cascade to yield pyruvate as co-substrate for L-tyrosine production Xiaolei Guo 1 & Weibin Wu 1 & Mingliang Zhang 1 & Licheng Wu 2 & Jianzhong Huang 1,3 Received: 19 May 2020 / Revised: 7 October 2020 / Accepted: 25 October 2020 # Springer-Verlag GmbH Germany, part of Springer Nature 2020
Abstract L-Tyrosine is a versatile compound used in the fine chemical, pharmaceutical, and functional food industries. Here, we report a bi-enzymatic cascade involving alanine racemase (ALR) and D-amino acid oxidase (DAAO) to produce pyruvate, as co-substrate for L-tyrosine production, from the cheap substrate L-alanine. The BpALR (ALR from Bacillus pseudofirmus) was used as a whole-cell biocatalyst, converting L-alanine to D, L-alanine. The FsDAAO (DAAO from Fusarium solani) was immobilized to oxidize the D-alanine generated in the first step to pyruvate. Both systems were combined as a continuous-flow reactor for maximized L-alanine-to-pyruvate conversion rates. The optimal parameters and appropriate conditions for FsDAAO immobilization were investigated. The pyruvate concentration of 86.6 g/L was achieved within 17 h. Subsequently, a whole-cell biocatalyst system for L-tyrosine production, catalyzed by the tyrosine phenol-lyase (TPL) from Erwinia herbicola (EhTPL), was developed, and a fed-batch approach was applied with phenol and the pyruvate produced with the ALR/DAAO system mentioned above. The concentration of phenol and pyruvate in the reactor should not exceed 7.5 g/L and 10 g/L, respectively. Significantly, the L-tyrosine concentration of 152.5 g/L was achieved within 10 h, demonstrating the great potential for highefficiency production of L-tyrosine through the approach we established in this paper.
Key points • A specific bioreactor system for pyruvate produced from L-alanine was developed • The appropriate condition for immobilization of FsDAAO was investigated • A fed-batch process was established to produce L-tyrosine with recombinant E. coli • The bi-enzymatic cascade was successfully used for L-tyrosine production at low cost Keywords L-tyrosine . Pyruvate . Bi-enzymatic cascade . Alanine racemase . D-amino acid oxidase . Tyrosine phenol-lyase
Introduction The aromatic amino acid L-tyrosine is a non-essential amino acid. It is required to make several compounds such as
* Jianzhong Huang [email protected] 1
College of Life Science, Fujian Normal University, Fuzhou 350117, Fujian, China
2
College of Life Science and Technology, State Key Laboratory of Agricultural Microbiology, Huazhong Agricultural University, Wuhan 430070, China
3
Engineering Research Center of Industrial Microbiology, Ministry of Education, Fujian Normal University, Fuzhou 350117, Fujian, China
hydroxytyrosol (Satoh et al. 2012), resveratrol (Zhao et al. 2018), caffeic acid (Rodrigues et al. 2015a), and curcuminoids (Rodrigues et al. 2015b) that are widely applied in the fine chemical, pharmaceutical, and functional food industries as adulterants or supplements. Previously
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