Increasing the bioflocculant production and identifying the effect of overexpressing epsB on the synthesis of polysaccharide and γ-PGA in Bacillus licheniformis

Abstract

BackgroundPolysaccharides and poly-γ-glutamic acid (γ-PGA) are biomacromolecules that have been reported as bioflocculants, and they exhibit high flocculating activity in many industrial applications. Bacillus licheniformis CGMCC 2876 can produce polysaccharide and γ-PGA bioflocculants under different culture conditions. Several key genes are involved in the metabolic pathway of polysaccharides in B. licheniformis, but the impacts of the regulation of these genes on the production of polysaccharide bioflocculants have not been illustrated completely. To increase the bioflocculant production and identify the correlation between the synthesis of polysaccharides and γ-PGA in B. licheniformis, a few key genes were investigated to explore their influence on the synthesis of the bioflocculants.ResultsOverexpressing epsB from the eps gene cluster not only improved the bioflocculant crude yield by 13.98% but also enhanced the flocculating activity by 117.92%. The composition of the bioflocculant from the epsB recombinant strain was 28.95% total sugar, 3.464% protein and 44.03% γ-PGA, while in the original strain, these components represented 53.67%, 3.246% and 34.13%, respectively. In combination with an analysis of the transcriptional levels of several key genes involved in γ-PGA synthesis in B. licheniformis, we inferred that epsB played a key role in the synthesis of both polysaccharide and γ-PGA. The bioflocculant production of the epsB recombinant strain was further evaluated during batch fermentation in a 2 L fermenter; the flocculating activity reached 9612.75 U/mL, and the bioflocculant yield reached 10.26 g/L after 72 h, representing increases of 224% and 36.62%, respectively, compared with the original strain. Moreover, we found that the tandem expression of phosphoglucomutase (pgcA) and UTP-glucose-1-phosphate uridylyltransferase (gtaB1) could enhance the crude yield of the bioflocculant by 20.77% and that the overexpression of epsA could enhance the bioflocculant yield by 23.70% compared with the original strain.ConclusionsThis study provides a new method to greatly increase the bioflocculant production in B. licheniformis, and it demonstrates the correlation between the biosynthesis of polysaccharide and γ-PGA during EPS fermentation by regulating the expression of EpsB.Electronic supplementary materialThe online version of this article (doi:10.1186/s12934-017-0775-9) contains supplementary material, which is available to authorized users.

Putative functions of EpsK in teichuronic acid synthesis and phosphate starvation in Bacillus licheniformis

“…In our previous study, we found that B. licheniformis CGMCC 2876 could produce polysaccharides and γ-PGA under different culture conditions [ 21 ]. EpsK was reported to play an important role in polysaccharide biosynthesis, however, its exact function has not been characterized [ 16 ].…”

“…After the fermentation process, the biopolymer was extracted and purified using the method described in a previous study [ 21 ]. The content of total carbohydrates in the biopolymer was detected using the phenol–sulfuric acid method.…”

Whole-Genome Sequencing and Potassium-Solubilizing Mechanism of Bacillus aryabhattai SK1-7

“…In this study, the polysaccharide content secreted by strain SK1-7 under an insoluble potassium source was higher than that under a soluble potassium source. The expression of the epsB gene was significantly upregulated with the addition of an insoluble potassium source, and it was upregulated 9.55 times at 10 h. epsB is the key gene regulating extracellular polysaccharide synthesis ( Elsholz et al, 2014 ; Liu et al, 2017 ). When cultured to the later growth of bacteria, it began to secrete a large amount of EPSs, which has a certain adhesion and can be used as an intermediate medium between potassium bacteria and mineral particles to bond them together to form a bacterial-mineral complex.…”

Genome analysis of a thermophilic exopolysaccharide-producing bacterium - Geobacillus sp. WSUCF1

“…The mutant with improved EPS production demonstrated that the specific activities of phosphoglucomutase (5.4.2.2), UDP-glucose pyrophosphorylase (2.7.7.9) and UDP-galactose-4-epimerase (5.1.3.2) were higher than those of the wild-type strain 49–51 , indicating potential targets for improvement of EPS production in strain WSUCF1 through homologous overexpression. The expression level of enzymes associated with GDP-mannose synthesis may also be important since multiple interventions during NDP-sugar synthesis are more likely to be efficient to promote EPS production 52,53 . As the second stage for EPS biosynthesis, GTs are responsible for the assembly of activated NDP-sugars to the polysaccharide chain.…”