Migration of glutamate decarboxylase by cold treatment on whole-cell biocatalyst triggered activity for 4-aminobutyric acid production in engineering Escherichia coli

• Published in: International journal of biological macromolecules 2021-11, Vol.190, p.113-119
• Main Authors: Xue, Chengfeng, Yi, Ying-Chen, Ng, I-Son
• Format: Article
• Language: English
• Subjects: 4-Aminobutyric acid; Biocatalysis; Cold Temperature; Cold treatment; Escherichia coli - genetics; Escherichia coli - ultrastructure; gamma-Aminobutyric Acid - biosynthesis; Genetic Engineering; Glutamate decarboxylase; Glutamate Decarboxylase - metabolism; Kinetics; Oxygen; Periplasm; Replication Origin - genetics; Whole-cell biocatalyst
• ISSN: 0141-8130; 1879-0003
• DOI: 10.1016/j.ijbiomac.2021.08.166

Glutamate decarboxylase B (GadB) from Escherichia coli, an intrinsic pyridoxal 5′-phosphate (PLP)-dependent enzyme has been employed for 4-aminobutyric acid (GABA) biosynthesis, which involves the glutamate import and GABA export via a transporter located in the inner membrane as rate determined step of whole-cell (WC) biotransformation. Herein, GadB was cloned and overexpressed in E. coli under a constitutive promoter in a high copy number plasmid, and 46.9 g/L GABA was produced. It was observed that GadB migrated to the periplasm when the WC were subjected to −20 °C cold treatment for 24 h prior to the biotransformation. Kinetic studies indicated that the enzymatic turnover rate of WC increased 2-fold after cold treatment, which was correlated with the migration rate of GadB, and up to 88.6% of GadB. The export or possible migration of GadB mitigated the rate-limiting step of WC biotransformation, and a 100% conversion of substrate to GABA was obtained. Finally, we launched a promising strategy for GABA production of 850 g/L from cost-effective monosodium glutamate (MSG) by using WC biocatalysts with 10-times recycling. •The constitutive Anderson promoter J23100 is effective for GadB expression.•The first attempt to explore GadB crossing to peri-plasma after cold treatment.•Cold treatment on whole-cell biocatalyst significantly increased enzyme activity.•Isothermal titration calorimetry revealed the interaction of GadB with co-factor PLP.•The highest 850 g/L GABA production was obtained by recycling whole-cell biocatalyst.