The Route of Sucrose Utilization by Streptococcus mutans Affects Intracellular Polysaccharide Metabolism

Streptococcus mutans converts extracellular sucrose (Suc) into exopolysaccharides (EPS) by glucosyl-transferase and fructosyl-transferase enzymes and internalizes Suc for fermentation through the phosphotransferase system (PTS). Here, we examined how altering the routes for sucrose utilization impacts intracellular polysaccharide [IPS; glycogen, (glg)] metabolism during carbohydrate starvation. Strain UA159 (WT), a mutant lacking all exo-enzymes for sucrose utilization (MMZ952), and a CcpA-deficient mutant (∆ccpA) were cultured with sucrose or a combination of glucose and fructose, followed by carbohydrate starvation. At baseline (0h), and after 4 and 24h of starvation, cells were evaluated for mRNA levels of the glg operon, IPS storage, glucose-1-phosphate (G1P) concentrations, viability, and PTS activities. A pH drop assay was performed in the absence of carbohydrates at the baseline to measure acid production. We observed glg operon activation in response to starvation (p<0.05) in all strains, however, such activation was significantly delayed and reduced in magnitude when EPS synthesis was involved (p<0.05). Enhanced acidification and greater G1P concentrations were observed in the sucrose-treated group, but mostly in strains capable of producing EPS (p<0.05). Importantly, only the WT exposed to sucrose was able to synthesize IPS during starvation. Contrary to CcpA-proficient strains, IPS was progressively degraded during starvation in ∆ccpA, which also showed increased glg operon expression and greater PTS activities at baseline. Therefore, sucrose metabolism by secreted enzymes affects the capacity of S. mutans in synthesizing IPS and converting it into organic acids, without necessarily inducing greater expression of the glg operon.

The Structural Characteristics and Biological Activities of Intracellular Polysaccharide Derived from Mutagenic Sanghuangporous sanghuang Strain

Sanghuangporous sanghuang is a rare medicinal fungus which contains polysaccharide as the main active substance and was used to treat gynecological diseases in ancient China. The intracellular polysaccharide yield of S. sanghuang was enhanced by the strain A130 which was screened from mutant strains via atmospheric and room temperature plasma (ARTP) mutagenesis. The objective of this research was to investigate the effects of ARTP mutagenesis on structural characteristics and biological activities of intracellular polysaccharides from S. sanghuang. Six intracellular polysaccharide components were obtained from S. sanghuang mycelia cultivated by the mutagenic strain (A130) and original strain (SH1), respectively. The results revealed that the yields of polysaccharide fractions A130-20, A130-50 and A130-70 isolated from the mutagenic strain fermentation mycelia were significantly higher than those of the original ones by 1.5-, 1.3- and 1.2-fold, and the clear physicochemical differences were found in polysaccharide fractions precipitated by 20% ethanol. A130-20 showed a relatively expanded branching chain with higher molecular weight and better in vitro macrophage activation activities and the IL-6, IL-1, and TNF-α production activities of macrophages were improved by stimulation of A130-20 from the mutagenic strain. This study demonstrates that ARTP is a novel and powerful tool to breed a high polysaccharide yield strain of S. sanghuang and may, therefore, contribute to the large-scale utilization of rare medicinal fungi.