期刊
BIOTECHNOLOGY AND BIOENGINEERING
卷 118, 期 7, 页码 2597-2608出版社
WILEY
DOI: 10.1002/bit.27780
关键词
Bacillus coagulans; dual‐ phase fed‐ batch fermentation; malic acid; metabolic and promoter engineering; reductive TCA pathway
资金
- Panjin Campus for Food Science and Technology Research Initiative, Dalian University of Technology
In this study, Bacillus coagulans was engineered into an efficient malic acid producer by establishing a reductive tricarboxylic acid pathway and optimizing promoters. Through screening and fine-tuning, the titer of malic acid was significantly improved, providing an effective approach for industrialized production using Bacillus coagulans.
Bacillus coagulans, a thermophilic facultative anaerobe, is a favorable chassis strain for the biosynthesis of desired products. In this study, B. coagulans was converted into an efficient malic acid producer by metabolic engineering and promoter engineering. Promoter mapping revealed that the endogenous promoter P-ldh was a tandem promoter. Accordingly, a promoter library was developed, covering a wide range of relative transcription efficiencies with small increments. A reductive tricarboxylic acid pathway was established in B. coagulans by introducing the genes encoding pyruvate carboxylase (pyc), malate dehydrogenase (mdh), and phosphoenolpyruvate carboxykinase (pckA). Five promoters of various strengths within the library were screened to fine-tune the expression of pyc to improve the biosynthesis of malic acid. In addition, genes involved in the competitive metabolic pathways were deleted to focus the substrate and energy flux toward malic acid. Dual-phase fed-batch fermentation was performed to increase the biomass of the strain, further improving the titer of malic acid to 25.5 g/L, with a conversion rate of 0.3 g/g glucose. Our study is a pioneer research using promoter engineering and genetically modified B. coagulans for the biosynthesis of malic acid, providing an effective approach for the industrialized production of desired products using B. coagulans.
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