Microbial production of poly-γ-glutamic acid (γ-PGA) from non-food raw materials is a promising alternative to food feedstocks-based biosynthesis. A superior cell factory of Bacillus amyloliquefaciens for the efficient synthesis of γ-PGA from crude glycerol was constructed through systematic metabolic engineering. Firstly, some phase-dependent promoters were screened from B. amyloliquefaciens, which can be used for fine regulation of subsequent metabolic pathways. Secondly, the glycerol utilization pathway and the γ-PGA synthesis pathway were co-optimized utilizing the above-screened promoters, which increased the titer of γ-PGA by 1.75-fold. Then, the titer of γ-PGA increased to 15.6 g/L by engineering transcription factors degU and blocking competitive pathways. Finally, combining these strategies with an optimized fermentation process, 26.4 g/L γ-PGA was obtained from crude glycerol as a single carbon source (a 3.72-fold improvement over the initial strain). Overall, these strategies will have great potential for synthesizing other products from crude glycerol in B. amyloliquefaciens.
Keywords: B. amyloliquefaciens; Crude glycerol; Modular metabolic engineering; Promoter; Transcription factors; γ-PGA.
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