Anti-α-glucosidase, Anti-proliferative and Anti-enterovirus 71 Activity of Secondary Metabolites Identified from Grifola Frondosa.

Grifola frondosa, an edible and medicinal resource, is widely used as functional foods worldwide. To explore bioactive compounds against α-glucosidase, human tumor cells and enterovirus 71 (EV71), eight compounds were isolated from G. frondosa by chromatographic column. Among the isolated compounds, heptadecanoic acid, uridine and adenosine exhibited potent inhibition activity against α-glucosidase, ergosterols and ergosterol-5,8-peroxide showed anti-proliferative activity on tumor cells, while ergosterol and methyl linoleate displayed inhibition against the replication of EV71. Also, to our knowledge, this is the first study to report that fatty acids isolated from G. frondosa show potent inhibition against α-glucosidase and EV71. Further molecular docking results revealed that the active compounds in G. frondosa form hydrogen bonding, hydrophobic interactive and π-stacking with the active sites on the surface of α-glucosidase, CASP3 and VP1 proteins, thus promoting the active compounds to combine with the target protein to form a stable complex, thus playing an antagonistic role. Our results could provide a new active compound and mode of action for G. frondosa to treat diabetes, cancer and EV71-infected patients.

Synergistic Antitumor Effect of Grifola frondose Polysaccharide-Protein Complex in Combination with Cyclophosphamide in H22 Tumor-Bearing Mice.

Hepatocellular carcinoma (HCC) is the most common type of liver malignancy and remains a global health threat. The objective of the current study was to determine whether the combination of a cold-water extracted polysaccharide-protein complex from Grifolia frondosa (GFG) and cyclophosphamide (CTX) could inhibit tumor growth by suppressing the expression of angiogenesis-related proteins in H22 tumor-bearing mice. The results showed that the inhibition rate of GFG combined with CTX on H22 tumors was 65.29%, which was significantly higher than that of GFG treatment alone (24.82%). GFG combined with CTX significantly increased the expression levels of vascular endothelial growth factor, basic fibroblast growth factor, matrix metalloproteinase 2, and matrix metalloproteinase 9. Additionally, thymus index, spleen index, natural killer (NK) cell activity, interferon-γ (IFN-γ), interleukin-1ß (IL-1ß), tumor necrosis factor-α (TNF-α) and interleukin-2 (IL-2) levels increased significantly after GFG treatment, especially after high-doses of GFG combined with CTX treatment (p < 0.05). The thymus index, spleen index, NK cell activity, IFN-γ, IL-1ß, TNF-α, and IL-2 levels were 1.90, 1.46, 1.30, 2.13, 1.64, 2.03, and 1.24 times of those treated with CTX alone. Thus, we proposed that GFG can alleviate the side effects of CTX by relieving the immunosuppressive effect, liver/renal injury, and oxidative stress. In conclusion, the combination of GFG and CTX for cancer treatment may be a promising strategy, and GFG is expected to be a potential adjuvant alternative for the treatment of HCC.

A Se-enriched Grifola frondosa polysaccharide induces macrophage activation by TLR4-mediated MAPK signaling pathway.

Se-polysaccharide (Se-GFP-22) from Se-enriched Grifola frondosa has double and cooperative activities of polysaccharide and Se. To delineate the underlying mechanism and signaling cascade involved in immune-stimulatory property of Se-GFP-22, the production of cellular mediators and key proteins in signaling pathway was examined. Results showed that Se-GFP-22 exhibited no cytotoxic and had a high capacity to promote macrophage phagocytosis, up-regulate interleukin-2 (IL-2), tumor necrosis factor-α (TNF-α), interferon-γ (IFN-γ), and nitric oxide (NO) productions, as well as the relative messenger RNA (mRNA) expressions. In Se-GFP-22-induced macrophages, intracellular superoxide dismutase (SOD) activity was significantly increased to protect cells from oxidative injury. However, Se-GFP-22 induced macrophage activation was suppressed when the toll-like receptor 4 (TLR4) signaling pathway was blocked by a specific TLR4 inhibitor. According to the western blot analysis and the use of specific inhibitors against the mitogen-activated protein kinases (MAPK) signaling pathway, we speculated that Se-GFP-22 activated RAW264.7 macrophages through the TLR4-mediated MAPK signaling pathway. This study provides a molecular basis for the potential of Se-GFP-22 as a novel immune-stimulatory agent.

Effect of low-intensity magnetic field on the growth and metabolite of Grifola frondosa in submerged fermentation and its possible mechanisms.

This study aimed to investigate the effects of low intensity alternating magnetic field on the submerged fermentation of Grifola frondosa, and its possible mechanism was also explored. Under the optimal shaking flask conditions, amino acids in mycelium with magnetic field treatment significantly increased, and the morphology of mycelium obviously changed. During the scale-up magnetic field-assisted fermentation, Mycelium biomass increased by 12%. The yield of polysaccharides and relative dissolved oxygen in the fermentation broth was higher than in the control group. Transcriptome sequencing results showed that the expression of genes related to amino acid metabolism increased significantly after magnetic treatment. In addition, magnetic field stimulation enhanced the mycelium biomass by upregulation the expression of genes related to cell repair and stress response. This study suggested that applying a magnetic field in submerged fermentation of G frondosa is an innovative approach to produce metabolites.

A cold-water polysaccharide-protein complex from Grifola frondosa exhibited antiproliferative activity via mitochondrial apoptotic and Fas/FasL pathways in HepG2 cells.

Grifola frondosa (G. frondosa) is widely known for its anti-tumor potential, which has been demonstrated by numerous scientific researches. In this study, two water soluble polysaccharide-protein complexes were extracted from G. frondosa at 4 °C (GFG-4) and 100 °C (GFG-100) and purified. Compared with GFG-100, GFG-4 had a higher protein content and molecular weight. The main monosaccharides of GFG-4 and GFG-100 were rhamnose, glucose, and galactose, with an approximate ratio of 3.00: 1.00: 0.86 and 2.85: 1.00: 0.94, respectively. The Fourier transform infrared spectra indicated that the two polysaccharide-protein complexes displayed characteristic functional groups of polysaccharides and proteins, and mainly contain pyranose ring with α-glycosidic linkage. Atomic force microscope images showed that both GFG-4 and GFG-100 exhibited straight chains, and GFG-4 possessed a relatively abundant fraction of branched chains. Intriguingly, GFG-4 showed a stronger antiproliferative activity against HepG2 cells than GFG-100. The mechanisms were further investigated by quantitative real-time PCR and western blot, it found that GFG-4 inhibited the proliferation of HepG2 cells mainly through the intrinsic activation of mitochondrial pathway and the Fas/FasL-mediated Caspase-8/-3 pathway. Conclusively, G. frondosa cold-water extracted polysaccharide-protein complexes could be used as a functional food for preventing or treating hepatocellular carcinoma.

Grifola frondosa Polysaccharide Ameliorates Early Diabetic Nephropathy by Suppressing the TLR4/NF-κB Pathway.

Grifola frondosa is a medicinal macro-fungus with a wide range of biological activities. Polysaccharides from Grifola frondosa (PGF) play a positive role in regulating blood glucose and alleviating kidney injury. Here, we investigated the exact mechanism of action by which PGF ameliorates diabetic nephropathy. Our results showed that PGF effectively improved glucose tolerance and insulin sensitivity in streptozocin (STZ)-induced DN mice. Additionally, administration of PGF also ameliorated renal function and inflammatory response in STZ-induced DN mice. Consistent with the in vitro results, the high glucose-induced inflammatory response and apoptosis of renal tubular epithelial cells were decreased by PGF treatment. Furthermore, PGF not only suppressed the expression of TLR4, but also more effectively protected the kidney and reduced the inflammatory response when TLR4 was inhibited. All these data revealed that PGF alleviates diabetic nephropathy by blocking the TLR4/NF-κB pathway.

Polysaccharides of Grifola frondosa ameliorate oxidative stress and hypercholesterolaemia in hamsters fed a high-fat, high-cholesterol diet.

OBJECTIVES: This study was to evaluate the antioxidant and anti-hypercholesterolaemia activities of Grifola frondosa in hamsters fed a high-fat, high-cholesterol (HFHC) diet. METHODS: G. frondosa, including fruiting bodies (FGF), fermented mycelia (MGF) and polysaccharides extracted from fruiting bodies (FPS), fermented mycelia (MIP) and fermented broth (BEP) were received intragastrically. Lipid profile and antioxidant status in the blood and liver of hamsters were assessed. KEY FINDINGS: FGF decreased weight gain, serum triglycerides and cholesterol and increased hepatic mRNA expression of cholesterol-7α-hydroxylase expression. FGF, MGF, FPS and MIP decreased the HFHC diet-increased area under the curve (AUC) of serum cholesterol. FGF and FPS further decreased AUC of serum triglycerides. When evaluating the redox status of erythrocytes, FPS and MIP increased non-protein sulfhydryl (NP-SH) groups, reduced glutathione (GSH) and catalase activity and FPS further increased GSH peroxidase activity. In the liver, MGF increased NP-SH groups and GSH and decreased triglycerides content. FPS, MIP and BEP decreased oxidized GSH and triglycerides content. Moreover, all treatments alleviated HFHC diet-increased LDL oxidation. CONCLUSIONS: Fruiting bodies of G. frondosa may improve hypercholesterolaemia via increased bile acid synthesis. Additionally, fermented biomass and polysaccharides of G. frondosa may have the potential to prevent hepatic lipid accumulation.

Fermentability of Maitake polysaccharides processed by various hydrothermal conditions and fermented with probiotic (Lactobacillus).

Maitake polysaccharides, after hydrothermal processing, were fermented with Lactobacillus acidophilus CCFM202 (L.A.) and Lactobacillus plantarum CCFM6392 (L.P.). The degradation of molecular weight of polysaccharides by hydrothermal processing under acidic conditions was obviously enhanced, which turned part of the water-insoluble-polysaccharides (WIP) into water-soluble-polysaccharides (WSPs). The pH value of water-soluble-polysaccharides (WSPs) and water-insoluble-polysaccharides (WIPs) were intensely dropped (4- 5) after 24 h fermentation. The optical density (O.D.) was increased (1.4- 2.3) due to bacterial growth, and short-chain fatty acids also followed this trend. LA-WSP predominantly produced acetic acid, 3- 4 folds to lactic acid, while LP-WIP groups produced dominant butyric acid (15- 17 folds). Hydrothermal processing induced the growth of L.A. and L.P., where the highest abundance was 2.5 × 104. From the Venn diagram, WSP-1 produced the most elevated metabolites (874). Therefore, experimental results show a significant impact on making WSPs fragments, whereas temperature and pH influence the WSPs degradation, withstand to higher fermentation efficacy.

Purification and characterization of Se-enriched Grifola frondosa glycoprotein, and evaluating its amelioration effect on As3+ -induced immune toxicity.

BACKGROUND: Selenium (Se)-enriched glycoproteins have been a research highlight for the role of both Se and glycoproteins in immunoregulation. Arsenic (As) is a toxicant that is potentially toxic to the immune function and consequently to human health. Several reports suggested that Se could reduce the toxicity of heavy metals. Moreover, more and more nutrients in food had been applied to relieve As-induced toxicity. Hence glycoproteins were isolated and purified from Se-enriched Grifola frondosa, and their preliminary characteristics as well as amelioration effect and mechanism on As3+ -induced immune toxicity were evaluated. RESULTS: Four factions, namely Se-GPr11 (electrophoresis analysis exhibited one band: 14.32 kDa), Se-GPr22 (two bands: 20.57 and 31.12 kDa), Se-GPr33 (three bands: 15.08, 20.57 and 32.78 kDa) and Se-GPr44 (three bands: 16.73, 32.78 and 42.46 kDa), were obtained from Se-enriched G. frondosa via DEAE-52 and Sephacryl S-400 column. In addition, Se-GPr11 and Se-GPr44 are ideal proteins that contain high amounts of almost all essential amino acids. Thereafter, the RAW264.7 macrophage model was adopted to estimate the effect of Se-GPr11 and Se-GPr44 on As3+ -induced immune toxicity. The results showed that the pre-intervention method was the best consequent and the potential mechanisms were, first, by improving the oxidative stress state (enhancing the activity of superoxide dismutase and glutathione peroxidase, decreasing the levels of reactive oxygen species and malondialdehyde); secondly, through nuclear factor-κB and mitogen-activated protein kinase-mediated upregulation cytokines (interleukin-2 and interferon-γ) secretion induced by As3+ . CONCLUSION: The results suggested Se-enriched G. frondosa may be a feasible supplement to improve health level of the As3+ pollution population. © 2021 Society of Chemical Industry.

Grifola frondosa may play an anti-obesity role by affecting intestinal microbiota to increase the production of short-chain fatty acids.

Background: Grifola frondosa (G. frondosa) is a fungus with good economic exploitation prospects of food and medicine homologation. This study aims to investigate the effects of G. frondosa powder suspension (GFPS) on the intestinal contents microbiota and the indexes related to oxidative stress and energy metabolism in mice, to provide new ideas for developing G. frondosa weight loss products. Methods: Twenty Kunming mice were randomly divided into control (CC), low-dose GFPS (CL), medium-dose GFPS (CM), and high-dose GFPS (CH) groups. The mice in CL, CM, and CH groups were intragastrically administered with 1.425 g/(kg·d), 2.85 g/(kg·d), and 5.735 g/(kg·d) GFPS, respectively. The mice in CC group were given the same dose of sterile water. After 8 weeks, liver and muscle related oxidative stress and energy metabolism indicators were detected, and the intestinal content microbiota of the mice was detected by 16S rRNA high-throughput sequencing. Results: After eight weeks of GFPS intervention, all mice lost weight. Compared with the CC group, lactate dehydrogenase (LDH) and malondialdehyde (MDA) contents in CL, CM, and CH groups were increased, while Succinate dehydrogenase (SDH) and Superoxide Dismutase (SOD) contents in the liver were decreased. The change trends of LDH and SDH in muscle were consistent with those in the liver. Among the above indexes, the change in CH is the most significant. The Chao1, ACE, Shannon, and Simpson index in CL, CM, and CH groups were increased. In the taxonomic composition, after the intervention with GFPS, the short-chain fatty acid (SCFA)-producing bacteria such as unclassified Muribaculaceae, Alloprevotella, and unclassified Lachnospiraceae increased. In linear discriminant analysis effect size (LEfSe) analysis, the characteristic bacteria in CC, CL, CM, and CH groups showed significant differences. In addition, some characteristic bacteria significantly correlated with related energy metabolism indicators. Conclusion: The preventive effect of G. frondosa on obesity is related to changing the structure of intestinal content microbiota and promoting the growth of SCFAs. While excessive intake of G. frondosa may not be conducive to the antioxidant capacity and energy metabolism.

The effects of quorum sensing molecule farnesol on the yield and activity of extracellular polysaccharide from Grifola frondosa in liquid fermentation.

A strategy by exogenous addition of quorum sensing molecule farnesol to improve the production, antioxidant activity and antitumor activity of extracellular polysaccharide (EPS) of Grifola frondosa by liquid fermentation was proposed in the study. The highest yield of EPS induced by farnesol was 1.25 g/L, which was 150% higher than that of the control. Four polysaccharides including EPS-C-0M, EPS-C-0.2M, EPS-F-0M and EPS-F-0.2M were extracted and purified under the conditions of control and farnesol respectively. The physicochemical properties, antioxidant activities and antitumor activities were studied. Their chemical composition differed in sugar, protein and uronic acid contents, and they were composed of six constituent monosaccharides with different ratios, with the average molecular weights of 1.12 × 103, 1.89 × 103, 1.41 × 103 and 2.02 × 103 kDa, respectively. They presented similar FT-IR spectra, but different surface morphology. Antioxidant experiments showed that they had strong scavenging activities on ABTS+, hydroxyl radical, O2- and DPPH radical. Antitumor experiments showed that they had strong inhibitory effects on human cervical cancer (HeLa) cells and human liver cancer cells (HepG2) cells. Among the four polysaccharides, EPS-F-0.2M showed the highest antioxidant and antitumor activities, indicating that farnesol could regulate the biological activity of EPS by affecting structure and properties. These results demonstrated that appropriate adjustment of culture conditions had potential application in the development of polysaccharides with high antioxidant and antitumor activity. It provided a new strategy to enhance the production and bioactivity of edible and medicinal fungal polysaccharides by using quorum sensing molecules.

Class I hydrophobins pretreatment stimulates PETase for monomers recycling of waste PETs.

Poly(ethylene terephthalate) hydrolase (PETase) from Ideonella sakaiensis 201-F6 was expressed and purified from Escherichia coli to hydrolyze poly(ethylene terephthalate) (PET) fibers waste for its monomers recycling. Hydrolysis carried out at pH 8 and 30 °C was found to be the optimal condition based on measured monomer mono(2-hydroxyethyl) terephthalate (MHET) and terephthalic acid (TPA) concentrations after 24 h reaction. The intermediate product bis(2-hydroxyethyl) terephthalate (BHET) was a good substrate for PETase because BHET released from PET hydrolysis was efficiently converted into MHET. Only a trace amount of MHET could be further hydrolyzed to TPA. Class I hydrophobins RolA from Aspergillus oryzae and HGFI from Grifola frondosa were expressed and purified from E. coli to pretreat PET surface for accelerating PETase hydrolysis against PET. The weight loss of hydrolyzed PET increased from approximately 18% to 34% after hydrophobins pretreatment. The releases of TPA and MHET from HGFI-pretreated PET were enhanced 48% and 62%, respectively. The selectivity (TPA/MHET ratio) of the hydrolysis reaction was approximately 0.5.

Effective adsorption of nisin on the surface of polystyrene using hydrophobin HGFI.

Herein, a new method was demonstrated for effective immobilization of the antibacterial peptide nisin on Grifola frondosa hydrophobin (HGFI), without the need of any additional complex reaction. Hydrophobin can self-assemble as a monolayer to form continuous negative-charged surfaces with enhanced wettability and biocompatibility. Adding nisin solution to such hydrophobin surface created antibacterial surfaces. The quantification analysis revealed that more nisin could be adsorbed on the HGFI-coated than to control polystyrene surfaces at different pH values. This suggested that electronic attraction and wettability may play important roles in this process. The transmission electron microscopy, atomic force microscopy and fourier transform infrared (FTIR) analysis indicated the adsorption mode of nisin on the HGFI film, i.e., hydrophobins served as an adhesive layer for binding charged peptides to interfaces. The antibacterial activity of the treated surface was investigated via counting, a nucleic acid release test, scanning electron microscopy, and biofilm detection. These results indicated the excellent antibacterial activity of nisin adsorbed on the HGFI-coated surfaces. The activity retention of adsorbed nisin was demonstrated by immersing the modified substrates in a flowed liquid condition.

A (1→6)-Branched (1→4)-ß-d-Glucan from Grifola frondosa Inhibits Lipopolysaccharide-Induced Cytokine Production in RAW264.7 Macrophages by Binding to TLR2 Rather than Dectin-1 or CR3 Receptors.

Mushroom polysaccharides including ß-glucans possess various health-promoting properties and are known to be the major bioactive constituents of Grifola frondosa (GF), which is a popular edible and medicinal mushroom. Dectin-1, a pattern-recognition receptor, is responsible for recognizing ß-glucans. In this study, parental RAW264.7 macrophages and Dectin-1-expressing RAW264.7 macrophages were used to investigate the anti-inflammatory activity and receptor involvement of the water-soluble polysaccharides from GF. Results indicated that the high molecular weight fraction of GF (GF70-F1; 1260 kDa) inhibited TNF-α and IL-6 production as well as NF-κB activation in lipopolysaccharide-induced macrophages. Chemical and enzymatic linkage analyses indicated that GF70-F1 mainly contained the known (1→3),(1→6)-ß-d-glucan and a polysaccharide not previously isolated from GF, a nondigestible glucan with a ß-(1→4)-linked backbone and ß-(1→6)-linked branches. The ability of GF70-F1 to inhibit cytokine production was not affected by the expression level of Dectin-1 in cells, and a similar inhibitory activity was observed after removing the (1→3),(1→6)-ß-d-glucan from GF70-F1. Blockade of Toll-like receptor 2 (TLR2) but not Dectin-1 or complement receptor 3 (CR3) attenuated the inhibitory activity of GF70-F1. The nondigestible (1→6)-branched (1→4)-ß-d-glucan in GF70-F1 may contribute to the anti-inflammatory activity via interacting with TLR2 rather than Dectin-1 or CR3 receptors.

Engineering Aspects in Production of Various Medicinal Mushrooms Biomass in Submerged Bioreactors.

Basidiomycetes of various species and their wide range of pharmaceuticaly interesting products in the past decades represents one of the most attractive groups of natural products in Asia and North America. Production of mushroom fruit bodies using farming technology is hardly covering the market. Development of comprehensive submerged technologies in stirred tank and air lift bioreactors are the most promising technologies for fast and large-amount cultivation of medicinal mushroom biomass and its pharmaceutically active products. Research in physiology, basic and applied studies in mushroom metabolism, process engineering aspects, and clinical studies in the past two decades represent a large cotribution to the development of this potential, which initiates the development of new drugs and some very attractive over-the-counter human and veterinary remedies. The current article is an overview of the most relevant engineering achievements in submerged cultivation of some medicinal mushrooms-Grifola frondosa, Trametes versicolor, Hericium erinaceus, and Cordyceps militaris-and some other species biomass production in bioreactors.

The Positive Effects of Grifola frondosa Heteropolysaccharide on NAFLD and Regulation of the Gut Microbiota.

: Non-alcoholic fatty liver disease (NAFLD) is a major public health problem in many countries. In this study, the ability of Grifola frondosa heteropolysaccharide (GFP) to ameliorate NAFLD was investigated in rats fed a high-fat diet (HFD). The molecular mechanisms modulating the expression of specific gene members related to lipid synthesis and conversion, cholesterol metabolism, and inflammation pathways were determined. The components of the intestinal microflora in rats were analyzed by high-throughput next-generation 16S rRNA gene sequencing. Supplementation with GFP significantly increased the proportions of Allobaculum, Bacteroides, and Bifidobacterium and decreased the proportions of Acetatifactor, Alistipes, Flavonifractor, Paraprevotella, and Oscillibacter. In addition, Alistipes, Flavonifractor, and Oscillibacter were shown to be significant cecal microbiota according to the Spearman's correlation test between the gut microbiota and biomedical assays (|r| > 0.7). Histological analysis and biomedical assays showed that GFP treatments could significantly protect against NAFLD. In addition, Alistipes, Flavonifractor, and Oscillibacter may play vital roles in the prevention of NAFLD. These results suggest that GFP could be used as a functional material to regulate the gut microbiota of NAFLD individuals.

Functions of a Glucan Synthase Gene GFGLS in Mycelial Growth and Polysaccharide Production of Grifola frondosa.

Glucan synthase (GLS) gene is known to be involved in the fungal biosynthesis of cell wall, differentiation, and growth. In the present study, a glucan synthase gene (GFGLS) in the edible mushroom Grifola frondosa with a full sequence of 5927 bp encoding a total of 1781 amino acids was cloned and characterized for the first time. GFGLSp is a membrane protein containing two large transmembrane domains connected with a hydrophilic cytoplasmic domain. With a constructed dual promoter RNA silencing vector pAN7-gfgls-dual, a GFGLS-silencing transformant iGFGLS-3 had the lowest GFGLS transcriptional expression level (26.1%) with a shorter length and thinner appearance of the mycelia, as well as decreased mycelial biomass and exo-polysaccharide production of 5.02 and 0.38 g/L, respectively. Further analysis indicated that GFGLS silence influenced slightly the monosaccharide compositions and ratios of mycelial and exo-polysaccharide. These findings suggest that GFGLS could affect mycelial growth and polysaccharide production by downregulating the glucan synthesis.

Anti-Diabetic and Anti-Nephritic Activities of Grifola frondosa Mycelium Polysaccharides in Diet-Streptozotocin-Induced Diabetic Rats Via Modulation on Oxidative Stress.

Grifola frondosa is an edible fungus with a variety of potential pharmacological activities. This study investigates the hypoglycemic, anti-diabetic nephritic, and antioxidant properties of G. frondosa polysaccharides in diet-streptozotocin-induced diabetic rats. After a 4-week treatment with 100 mg/kg of metformin and 200 mg/kg of one of four different G. frondosa polysaccharide mixtures (especially GFPS3 and GFPS4), diabetic rats had enhanced body weight and suppressed plasma glucose, indicating the hypoglycemic activities of the G. frondosa polysaccharides. G. frondosa polysaccharides regulated the level of serum creatinine, blood urea nitrogen, N-acetyl-ß-D-glucosaminidase, and albuminuria; inhibited the serum levels of interleukin (IL)-2, IL-6, and TNF-α; and enhanced the serum levels of matrix metalloproteinase 9 and interferon-α, confirming their anti-diabetic nephritic activities. G. frondosa polysaccharides ameliorated the pathological alterations in the kidneys of diabetic rats. Moreover, G. frondosa polysaccharides modulated the serum levels of oxidant factors such as superoxide dismutase, glutathione peroxidase, catalase, malondialdehyde, and reactive oxygen species, revealing their antioxidant properties. Furthermore, the administration of G. frondosa polysaccharides inhibited nuclear factor kappa B activities in the serum and kidneys. All of the data revealed that the activation of nuclear factor kappa B plays a central role in G. frondosa polysaccharide-mediated anti-diabetic and anti-nephritic activities.

Improved mycelia and polysaccharide production of Grifola frondosa by controlling morphology with microparticle Talc.

BACKGROUND: Mushroom showed pellet, clump and/or filamentous mycelial morphologies during submerged fermentation. Addition of microparticles including Talc (magnesium silicate), aluminum oxide and titanium oxide could control mycelial morphologies to improve mycelia growth and secondary metabolites production. Here, effect of microparticle Talc (45 µm) addition on the mycelial morphology, fermentation performance, monosaccharide compositions of polysaccharides and enzymes activities associated with polysaccharide synthesis in G. frondosa was well investigated to find a clue of the relationship between polysaccharide biosynthesis and morphological changes. RESULTS: Addition of Talc decreased the diameter of the pellets and increased the percentage of S-fraction mycelia. Talc gave the maximum mycelial biomass of 19.25 g/L and exo-polysaccharide of 3.12 g/L at 6.0 g/L of Talc, and mycelial polysaccharide of 0.24 g/g at 3.0 g/L of Talc. Talc altered the monosaccharide compositions/percentages in G. frondosa mycelial polysaccharide with highest mannose percentage of 62.76 % and lowest glucose percentage of 15.22 % followed with the corresponding changes of polysaccharide-synthesis associated enzymes including lowest UDP-glucose pyrophosphorylase (UGP) activity of 91.18 mU/mg and highest UDP-glucose dehydrogenase (UGDG) and GDP-mannose pyrophosphorylase (GMPPB) activities of 81.45 mU/mg and 93.15 mU/mg. CONCLUSION: Our findings revealed that the presence of Talc significantly changed the polysaccharide production and sugar compositions/percentages in mycelial and exo-polysaccharides by affecting mycelial morphology and polysaccharide-biosynthesis related enzymes activities of G. frondosa.

A mutant of hydrophobin HGFI tuning the self-assembly behaviour and biosurfactant activity.

Hydrophobins are a series of low molecular weight proteins produced by filamentous fungi that play an important role in fungal growth. They have a globular structure and possess a unique hydrophobic patch on their surface that makes them amphiphilic, making them among the most surface-active proteins. Herein, the surface charge properties of HGFI, a class I hydrophobin from Grifola frondosa, were altered by replacing the negatively charged Glu24 with a positively charged Lys to generate the ME24 mutant. Pichia pastoris GS115 was used for recombinant expression of the ME24 mutant, which was purified by a two-step procedure. The function of the mutated residue in HGFI self-assembly was investigated. Reverse-phase high-performance liquid chromatography analysis revealed that the polarity of ME24 was enhanced compared with HGFI. Circular dichroism, thioflavin T assay, water contact angle and atomic force microscopy indicated that Glu24 participates in rodlet formation. Water solubility detection and dynamic light scattering showed that Glu24 affects the assembled state of HGFI in aqueous solution. The behaviour of the mutant in an emulsion, in the dispersion of insoluble materials and in large-scaled protein production suggests the functions of hydrophobins can be tuned for new applications.