Enhanced Low Molecular Weight Poly-γ-Glutamic Acid Production in Recombinant Bacillus subtilis 1A751 with Zinc Ion

Poly-γ-glutamic acid (γ-PGA) is a novel biodegradable polyamide material. Microbial fermentation is the only way to produce γ-PGA, but the molecular weight of γ-PGA varied depending on different strains and culture conditions used. The molecular weight of γ-PGA is a main factor affecting the utilization of γ-PGA. It is urgent to find an efficient way to prepare γ-PGA with specific molecular weight, especially low molecular weight. Bacillus subtilis ECUST is a glutamate-dependent strain that produces γ-PGA. In this study, a recombinant B. subtilis harboring the γ-PGA synthase gene cluster pgsBCAE of our preciously identified γ-PGA-producing B. subtilis ECUST was constructed. Assay of γ-PGA contents and properties showed that recombinant B. subtilis 1A751-pBNS2-pgsBCAE obtained the ability to synthesize γ-PGA with low molecular weight (about 10 kDa). The excessive addition of glutamate inhibited the γ-PGA synthesis, while the addition of Zn²⁺ could promote the synthesis of γ-PGA by increasing the transcription of pgsB but had no effect on the molecular weight of synthesized γ-PGA. Under optimized conditions, γ-PGA produced by recombinant B. subtilis 1A751-pBNS2-pgsBCAE increased from initial 0.54 g/L to 3.9 g/L, and the glutamate conversion rate reached 78%. Recombinant B. subtilis 1A751-pBNS2-pgsBCAE has the potential for efficient preparation of low molecular weight γ-PGA.