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Enhanced Production of Poly-γ-glutamic acid by Overexpression of the Global Anaerobic Regulator Fnr in Bacillus licheniformis WX-02

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Abstract

Poly-γ-glutamic acid is a multi-functional biopolymer with various applications. ATP supply plays an important role in poly-γ-glutamic acid (γ-PGA) synthesis. Global anaerobic regulator Fnr plays a key role in anaerobic adaptation and nitrate respiration, which might affect ATP generation during γ-PGA synthesis. In this study, we have improved γ-PGA production by overexpression of Fnr in Bacillus licheniformis WX-02. First, the gene fnr was knocked out in WX-02, and the γ-PGA yields have no significant differences between WX-02 and the fnr-deficient strain WXΔfnr in the medium without nitrate (BFC medium). However, the γ-PGA yield of 8.95 g/L, which was produced by WXΔfnr in the medium with nitrate addition (BFCN medium), decreased by 74% compared to WX-02 (34.53 g/L). Then, the fnr complementation strain WXΔfnr/pHY-fnr restored the γ-PGA synthesis capability, and γ-PGA yield was increased by 13% in the Fnr overexpression strain WX/pHY-fnr (39.96 g/L) in BFCN medium, compared to WX/pHY300 (35.41 g/L). Furthermore, the transcriptional levels of narK, narG, and hmp were increased by 5.41-, 4.93-, and 3.93-fold in WX/pHY-fnr, respectively, which led to the increases of nitrate consumption rate and ATP supply for γ-PGA synthesis. Collectively, Fnr affects γ-PGA synthesis mainly through manipulating the expression level of nitrate metabolism, and this study provides a novel strategy to improve γ-PGA production by overexpression of Fnr.

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Funding

This work was supported by the National Program on Key Basic Research Project (973 Program, No. 2015CB150505) and the Science and Technology Program of Wuhan (20160201010086).

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Authors and Affiliations

  1. Environmental Microbial Technology Center of Hubei Province, Hubei Collaborative Innovation Center for Green Transformation of Bio-Resources, College of Life Sciences, Hubei University, Wuhan, 430062, China

    Dongbo Cai, Shiying Hu, Yaozhong Chen, Li Liu, Shihui Yang, Xin Ma & Shouwen Chen

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  1. Dongbo Cai
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  2. Shiying Hu
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  3. Yaozhong Chen
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  4. Li Liu
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Contributions

D Cai and S Chen designed the study. D Cai carried out the molecular biology studies and construction of engineering strains. D Cai, S Hu, Y Chen and L Liu carried out the fermentation studies. D Cai, S Yang, X, Ma and S Chen analyzed the data and wrote the manuscript. All authors read and approved the final manuscript.

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Correspondence to Xin Ma or Shouwen Chen.

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The authors declare that they have no conflict of interest.

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All the primers sequences for RT-qPCR (Table S1) were supplied as the Supplementary Materials. This information is available free of charge via the Internet http://www.springer.com/chemistry/biotechnology/journal/12010.

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Cai, D., Hu, S., Chen, Y. et al. Enhanced Production of Poly-γ-glutamic acid by Overexpression of the Global Anaerobic Regulator Fnr in Bacillus licheniformis WX-02. Appl Biochem Biotechnol 185, 958–970 (2018). https://doi.org/10.1007/s12010-018-2693-7

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