A water-soluble mycelium polysaccharide from Monascus pilosus: Extraction, structural characterization, immunomodulatory effect and yield enhanced by overexpression of UGE gene

Mycelium polysaccharide (MPP) from Monascus pilosus with the compositions of glucose, galactose, mannose, glucosamine hydrochloride, rhamnose and arabinose, was obtained using alkaline extracting, and subsequently three purified components (MPP-0, MPP-0.1 and MPP-0.3) were separated. The purity and extraction volume of the MPP-0.1 fraction surpassed those of the other two groups, thus warranting its selection for subsequent experimental investigations. The sample MPP-0.1, with an average molecular weight of 3.7776 x 10(4) Da, exhibited exceptional thermal stability up to 170 degrees C. The main glycosidic linkage pattern of MPP-0.1 was structured as ->[4)-alpha-D-Glcp-(1](6) -> 4)-alpha-D-Glcp-(1 -> [2)-alpha-D-Manp-(1](5) -> 2)-alpha-D-Manp-(1 -> 5)-beta-D-Galf-(1 -> 3)-beta-D-Galf (1 -> 3)-beta-D-Galf-(1 -> 3)-beta-D-Galf-(1 ->, and branched Glcp, Manp, Galf fragments were connected with the main chain through -> 4, 6)-alpha-D-Glcp-(1 ->, -> 2, 6)-alpha-D-Manp-(1 -> and -> 3, 6)-beta-D-Galf-(1 ->. Besides, the up-regulated levels of Nitric oxide (NO), Tumor necrosis factor-alpha (TNF-alpha), Interleukin-6 (IL-6), Interleukin-1 beta (IL-1 beta) and other pro-inflammatory cytokines along with increased phagocytic activity revealed that MPP-0.1 has significant immunomodulatory effect, and can significantly enhance the proliferation and activation of RAW264.7 cells. Finally, the gene UGE (UDP-glucose 4-epimerase) was overexpressed in M. pilosus to increase the MPP production. Results showed that the biomass of the recombinant strain exhibited a remarkable increase of approximately 62.56 +/- 1.50 % compared to that of the parental strain, and the extraction yield of MPP increased significantly by 83.19 +/- 4.56 %.