Inactivation of the Levansucrase Gene in Paenibacillus polymyxa DSM 365 Diminishes Exopolysaccharide Biosynthesis during 2,3-Butanediol Fermentation
May 06, 2020 | Biotechnology
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The formation of exopolysaccharides (EPSs) during 2,3-butanediol (2,3-BD) fermentation by Paenibacillus polymyxa increases medium viscosity, which in turn presents considerable technical and economic challenges to 2,3-BD downstream processing. To eliminate EPS production during 2,3-BD fermentation, we used homologous recombination to disable the EPS biosynthetic pathway in P. polymyxa. The gene which encodes levansucrase, the major enzyme responsible for EPS biosynthesis in P. polymyxa, was successfully disrupted. The P. polymyxa levansucrase null mutant produced 2.5 ± 0.1 and 1.2 ± 0.2 g/liter EPS on sucrose and glucose, respectively, whereas the wild type produced 21.7 ± 2.5 and 3.1 ± 0.0 g/liter EPS on the same substrates, respectively. These levels of EPS translate to 8.7- and 2.6-fold decreases in EPS formation by the levansucrase null mutant on sucrose and glucose, respectively, relative to that by the wild type, with no significant reduction in 2,3-BD production. Inactivation of EPS biosynthesis led to a considerable increase in growth. On glucose and sucrose, the cell biomass of the levansucrase null mutant (8.1 ± 0.8 and 6.5 ± 0.3 g/liter, respectively) increased 1.4-fold compared to that of the wild type (6.0 ± 0.1 and 4.6 ± 0.3 g/liter, respectively) grown on the same substrates. Evaluation of the genetic stability of the levansucrase null mutant showed that it remained genetically stable over fifty generations, with no observable decrease in growth or 2,3-BD formation, with or without antibiotic supplementation. Hence, the P. polymyxa levansucrase null mutant has potential for use as an industrial biocatalyst for a cost-effective large-scale 2,3-BD fermentation process devoid of EPS-related challenges.
IMPORTANCE Given the current barrage of attention and research investments toward the production of next-generation fuels and chemicals, of which 2,3-butanediol (2,3-BD) produced by nonpathogenic... More of this in the AEM: Applied and Environmental Microbiology Blog.
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