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Metabolic Engineering of Central Carbon Metabolism of Bacillus licheniformis for Enhanced Production of Poly-γ-glutamic Acid

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Abstract

Poly-γ-glutamic acid (γ-PGA) is an anionic polymer with wide-ranging applications in the areas of medicine, light chemical industry, wastewater treatment, and agriculture. However, the production cost of γ-PGA is high for the requirement of adding the expensive precursor L-glutamic acid during fermentation, which hinders its widespread application. In this study, in order to improve γ-PGA yield, central carbon metabolism was engineered to enhance the carbon flux of tricarboxylic acid (TCA) cycle and glutamic acid synthesis in a γ-PGA production strain Bacillus licheniformis WX-02. Firstly, pyruvate dehydrogenase (PdhABCD) and citrate synthase (CitA) were overexpressed to strengthen the flux of pyruvate into TCA cycle, resulting in 34.93% and 11.14% increase of γ-PGA yield in B. licheniformis WX-02, respectively. Secondly, the carbon flux to glyoxylate shunt was rewired via varying the expression of isocitrate lyase (AceA), and a 23.24% increase of γ-PGA yield was obtained in AceA down-regulated strain WXPbacAaceBA. Thirdly, deletion of pyruvate formate-lyase gene pflB led to a 30.70% increase of γ-PGA yield. Finally, combinatorial metabolic engineering was applied, and γ-PGA titer was enhanced to 12.02 g/L via overexpressing pdhABCD and citA, repressing aceA, and deleting pflB, with a 69.30% improvement compared to WX-02. Collectively, metabolic engineering of central carbon metabolism is an effective strategy for enhanced γ-PGA production in B. licheniformis, and this research provided a promising strain for industrial production of γ-PGA.

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Funding

This work was supported by the Educational Research Projects for Young and Middle-Aged Teachers in Fujian Province (No. JAT190788) and Fujian Provincial Key Laboratory of Eco-Industrial Green Technology (No. WYKF-GCT2020-3).

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Authors and Affiliations

  1. Fujian Provincial Key Laboratory of Eco-Industrial Green Technology, College of Ecological and Resource Engineering, Wuyi University, Wuyishan, 354300, People’s Republic of China

    Bichan Li & Shouwen Chen

  2. State Key Laboratory of Biocatalysis and Enzyme Engineering, Environmental Microbial Technology Center of Hubei Province, College of Life Sciences, Hubei University, Wuhan, 430062, People’s Republic of China

    Dongbo Cai & Shouwen Chen

  3. State Key Laboratory of Agricultural Microbiology, College of Life Science and Technology, Huazhong Agricultural University, Wuhan, 430070, Hubei, People’s Republic of China

    Bichan Li

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  1. Bichan Li
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  2. Dongbo Cai
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Contributions

B Li, D Cai, and S Chen designed the study. B Li carried out the molecular biology studies, construction of engineering strains, and the fermentation studies. B Li, D Cai, and S Chen analyzed the data and wrote the manuscript. All authors read and approved the final manuscript.

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Correspondence to Shouwen Chen.

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Li, B., Cai, D. & Chen, S. Metabolic Engineering of Central Carbon Metabolism of Bacillus licheniformis for Enhanced Production of Poly-γ-glutamic Acid. Appl Biochem Biotechnol 193, 3540–3552 (2021). https://doi.org/10.1007/s12010-021-03619-4

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