Part:BBa_K2963009

capB*CA - encoding poly-γ-glutamic acid synthetase

The gene cluster capB*CA encode an multi-enzyme complex which can polymorize D/L glutamic acid to synthesize γ-PGA.

Usage and Biology

The BCA genes from Bacillus sp., encoding a polyglutamate synthetase located on the cell membrane, is capable of polymerizing glutamic acid to produce γ-PGA. In Bacillus licheniformis, BCA are called capBCA. We used the mutant B* gene of B gene. This part is used for producing L-glutamate-rich γ-PGA.

Characterization

We used NMR to detect γ-PGA and HPLC to analyze L- glutamate ratio of γ-PGA we have produce. The results as follow show that we have successfully produced L-glutamate-rich γ-PGA.

By the NMR detection of the fermentation product, the specific hydrogen peaks a, b, and c on the γ-amide bond on the γ-polyglutamic acid could be detected at the corresponding time points. It is indicated that the capB*CA genes from Bacillus licheniformis heterologously express in Corynebacterium glutamicum, and the target product γ-PGA is successfully produced.

We used HPLC to detect the L-glutamate monomer ratio of the γ-PGA we have produced. The results show that the L-glutamic acid monomer ratio reaches more than 90%. This part is working and we have produced L-glutamate-rich γ-PGA.

References

1. Xu P, Vansiri A, Bhan N, et al. ePathBrick: a synthetic biology platform for engineering metabolic pathways in E. coli[J]. ACS Synthetic Biology, 2012, 1(7): 256-266.

2. Peng Yingyun. Study on the production, synthesis mechanism and antifreeze of γ-polyglutamic acid. Diss. Jiangnan University, 2015.

3. Sung M H, Park C, Kim C J, et al. Natural and edible biopolymer poly-gamma-glutamic acid: synthesis, production, and applications [J]. Chemical Record, 2005, 5(6): 352-366.