Coculture Corynebacterium glutamicum and Bacillus licheniformis for producing poly-γ-glutamic acid from glucose.

Poly-γ-glutamic acid (γ-PGA) has diverse applications from cosmetic to drug delivery. The production of γ-PGA primarily relies on microbial fermentation using Bacillus spp. supplemented with L-glutamate supplementation. However, the high cost of L-glutamate supplementation limits industrial production. This study aimed to achieve direct γ-PGA production from glucose using a Corynebacterium glutamicum-Bacillus licheniformis coculture system. To create such a coculture system, we utilized B. licheniformis ATCC 9945a, a natural L-glutamate-dependent γ-PGA producing strain, and C. glutamicum F343, which exhibited an excellent capacity to produce L-glutamate from glucose. B. licheniformis ATCC 9945a grew well and produced small amounts of γ-PGA in the medium of C. glutamicum F343. Subsequently, B. licheniformis ATCC 9945a was cultured using the supernatant collected from the C. glutamicum F343 fermentation broth to investigate its effect on the fermentation profile. It was found that B. licheniformis ATCC 9945a produced more γ-PGA in the supernatant compared to when exogenously supplemented with L-glutamate. Moreover, nine intracellular metabolites were discovered to be strongly connected to γ-PGA synthesis by UPLC-MS. Finally, the coculture of C. glutamicum F343 and B. licheniformis ATCC 9945a to produce γ-PGA was conducted. We successfully achieved direct γ-PGA production from glucose under optimal conditions, including an inoculation time of 4 h for B. licheniformis after C. glutamicum inoculation, a 75% inoculum ratio of C. glutamicum, and a total inoculum size of 10% culture volume. The coculture system produced 12.49 g/L of γ-PGA in a shake flask and 22.7 g/L in a 5-L fermentor. KEY POINTS: • C. glutamicum F343 could produce L-glutamate from glucose as a precursor for PGA synthesis by B. licheniformis ATCC 9945a. • The C. glutamicum-B. licheniformis coculture system could produce γ-PGA up to 22.7 g/L. • Nine intracellular metabolites demonstrated a remarkable influence on γ-PGA synesis by UPLC-MS and metabolite profiling.