Phosphoglucose Isomerase Activity Research Articles

Effect of inoculation with arbuscular mycorrhizal fungi on the energy metabolism of selenite-rich amaranth.

A series of pot trials were undertaken to examine the impact of four arbuscular mycorrhizal fungi (AMF), namely Glomus mosseae (G.m), Glomus etunicatum (G.e), Corymbiglomus tortuosum (C.t), and the combined application of Glomus etunicatum and Corymbiglomus tortuosum (G.e + C.t), on the energy metabolism of amaranth plants grown in soil enriched with selenite at a concentration of 0.5 mg kg-1 . The inoculation of four AMFs resulted in an increase in both amaranth biomass and selenium (Se) content in leaves. The activities of phosphoglucose isomerase (PGI) and glucose-6-phosphate dehydrogenase + 6-phosphogluconate dehydrogenase were observed to decrease when AMFs were inoculated, as compared with the absence of AMF inoculation. The inoculation with G.m, C.t, and G.e + C.t resulted in an increase in succinate dehydrogenase activity; however, the inoculation with G.m, G.e, and G.e + C.t led to an increase in ascorbate oxidase activity. Furthermore, the inoculation of all four AMFs resulted in an increase in cytochrome c oxidase activity and the concentrations of oxidized coenzyme I (NAD) and reduced coenzyme I (NADH). The polyphenol oxidase activity of amaranth plants increased when inoculated with G.m and G.e, whereas it decreased when inoculated with C.t and G.e + C.t. Furthermore, the application of all four AMF treatments resulted in a reduction in adenosine triphosphate (ATP) levels and energy charge. It was worth mentioning that there was a clear inverse relationship between the energy charge and the biomass, Se concentration in the leaves. The findings presented in this research indicated that AMF may have an impact on energy metabolism and ultimately the biomass of amaranth by influencing the uptake of Se.

PRMT5 regulates the polysaccharide content by controlling the splicing of thaumatin-like protein in Ganoderma lucidum.

In the current study, the silencing of protein arginine methyltransferase 5 (PRMT5) decreased the polysaccharide content of Ganoderma lucidum compared to that of the wild-type (WT) strain. Furthermore, RNA-seq screening showed that the glycan degradation process-related gene thaumatin-like protein (TLP) was alternatively spliced in WT strains via retained introns, leading to the production of a longer TLP1 isoform and a shorter TLP2 isoform; however, only the TLP2 isoform was observed in PRMT5i strains. Experiments examining the polysaccharide content of the TLP silencing, TLP1 overexpression (OE-TLP1), and TLP2 overexpression (OE-TLP2) transformants revealed that TLP2 plays a more important role than TLP1 in polysaccharide degradation. Through a combination of yeast two-hybrid, bimolecular fluorescence complementation and surface plasmon resonance assays, we found that TLP2 directly physically interacted with phosphoglucose isomerase (PGI), a key enzyme in polysaccharide synthesis, and thereby increased PGI activity. However, TLP1 failed to interact with PGI, and PGI activity was not affected. Further inspection showed that the polysaccharide content was decreased in the OE-TLP2 strains but not significantly changed in the OE-TLP1 strains compared with that in the WT strains. In addition, the polysaccharide content of the PRMT5-TLP-cosilenced strains was not significantly different from that of the WT strains. These results demonstrate that PRMT5 modulates TLP processing of pre-mRNA transcripts and thereby decreases the polysaccharide content.IMPORTANCEPRMT5 contributes to secondary metabolite biosynthesis in Ganoderma lucidum. However, the mechanism through which PRMT5 regulates the biosynthesis of secondary metabolites remains unclear. In the current study, PRMT5 silencing led to a significant decrease in the biosynthesis of polysaccharides from G. lucidum through the action of the alternative splicing of TLP. A shorter TLP2 isoform can directly bind to PGI and regulated polysaccharide biosynthesis. These results suggest that PRMT5 enhances PGI activity by regulating TLP binding to PGI. The results of the current study reveal a novel target gene for PRMT5-mediated alternative splicing and provide a reference for the identification of PRMT5 regulatory target genes.

Effects of phthalate and bisphenol plasticizers on the activity of glycolytic enzymes of the moth Spodoptera littoralis

Environmental plastic pollution has significantly increased in the recent decades, and severely impacts economies, human and biodiversity health. Plastics are made of several chemical additives, including bisphenol and phthalate plasticizers such as bisphenol A (BPA) and Di(2-ethylhexyl)phthalate (DEHP). In some animal species, both BPA and DEHP are known as endocrine disruptor compounds, and can alter physiological and metabolic homeostasis, reproduction, development and/or behavior. To date, the impacts of BPA and DEHP have mainly focused on vertebrates, and to a lesser extent, on aquatic invertebrates. Yet, the few studies which examined the effects of DEHP on terrestrial insects also revealed the impacts this pollutant can have on development, hormone titrations, and metabolic profiles. In particular, it has been hypothesized in the Egyptian cotton leafworm Spodoptera littoralis that the observed metabolic alterations could result from the energetic costs necessary for DEHP detoxification or to the dysregulation of hormonally-controlled enzymatic activities. To get additional insights into the physiological effects of bisphenol and phthalate plasticizers on the moth S. littoralis, larvae were fed with food contaminated by BPA, DEHP, or the mixture of both compounds. Then, activities of four glycolytic enzymes, hexokinase, phosphoglucose isomerase, phosphofructokinase, and pyruvate kinase were measured. BPA and/or DEHP had no effects on the activities of phosphofructokinase and pyruvate kinase. Conversely, BPA-contaminated larvae were characterized by a 1.9-fold increase in phosphoglucose isomerase activity, and BPA + DEHP-fed larvae had highly variable hexokinase activity. Overall, since no disruption of glycolytic enzyme was observed in DEHP-contaminated larvae, our work tended to demonstrate that exposure to bisphenol and DEHP increased the amount of oxidative stress experienced.

Enhanced Effects of Iron on Mycelial Growth, Metabolism and In Vitro Antioxidant Activity of Polysaccharides from Lentinula edodes.

The polysaccharides found in Lentinula edodes have a variety of medicinal properties, such as anti-tumor and anti-viral effects, but their content in L. edodes sporophores is very low. In this study, Fe2+ was added to the liquid fermentation medium of L. edodes to analyze its effects on mycelial growth, polysaccharide and enzyme production, gene expression, and the activities of enzymes involved in polysaccharide biosynthesis, and in vitro antioxidation of polysaccharides. The results showed that when 200 mg/L of Fe2+ was added, with 7 days of shaking at 150 rpm and 3 days of static culture, the biomass reached its highest value (0.28 mg/50 mL) 50 days after the addition of Fe2+. Besides, Fe2+ addition also enhanced intracellular polysaccharide (IPS) and exopolysaccharide (EPS) productions, the levels of which were 2.98- and 1.79-fold higher than the control. The activities of the enzymes involved in polysaccharides biosynthesis, including phosphoglucomutase (PGM), phosphoglucose isomerase (PGI), and UDPG-pyrophosphorylase (UGP) were also increased under Fe2+ addition. Maximum PGI activity reached 1525.20 U/mg 30 days after Fe2+ addition, whereas PGM and UGP activities reached 3607.05 U/mg and 3823.27 U/mg 60 days after Fe2+ addition, respectively. The Pearson correlation coefficient showed a strong correlation (p < 0.01) between IPS production and PGM and UGP activities. The corresponding coding genes of the three enzymes were also upregulated. When evaluating the in vitro antioxidant activities of polysaccharides, EPS from all Fe2+-treated cultures exhibited significantly better capacity (p < 0.05) for scavenging -OH radicals. The results of the two-way ANOVA indicated that the abilities of polysaccharides to scavenge O2− radicals were significantly (p < 0.01) affected by Fe2+ concentration and incubation time. These results indicated that the addition of iron provided a good way to achieve desirable biomass, polysaccharide production, and the in vitro antioxidation of polysaccharides from L. edodes.

A structural basis for the functional differences between the cytosolic and plastid phosphoglucose isomerase isozymes.

Phosphoglucose isomerase (PGI) catalyzes the interconversion between glucose-6-phosphate (G6P) and fructose-6-phosphate (F6P), thereby regulating sucrose synthesis in plant cells. In general, plants contain a pair of PGI isozymes located in two distinct compartments of the cell (cytosol and plastid) with differences in both the primary structure and the higher-order structure. Previously, we showed that the activity of cytosolic PGI (PGIc) is more robust (activity, thermal stability, substrate turnover rate, etc.) than that of the plastid counterpart (PGIp) in multiple organisms, including wheat, rice, and Arabidopsis. The crystal structures of apoTaPGIc (an isotype cytosol PGIc in Triticum aestivum), TaPGIc-G6P complex, and apoTaPGIp (an isotype plastid PGIp in Triticum aestivum) were first solved in higher plants, especially in crops. In this study, we detailed the structural characteristics related to the biochemical properties and functions of TaPGIs in different plant organelles. We found that the C-terminal domains (CTDs) of TaPGIc and TaPGIp are very different, which affects the stability of the dimerized enzyme, and that Lys213TaPGIc/Lys193TaPGIp and its surrounding residues at the binding pocket gateway may participate in the entrance and exit of substrates. Our findings provide a good example illuminating the evolution of proteins from primary to higher structures as a result of physical barriers and adaptation to the biochemical environment.

Anthocyanin extract from Lycium ruthenicum enhanced production of biomass and polysaccharides during submerged fermentation of Agaricus bitorquis (Qu\xe9l.) Sacc. Chaidam

Agaricus bitorquis (Quél.) Sacc. Chaidam (ABSC) is a wild edible fungus uniquely found in the Tibet Plateau. ABSC is rich in polysaccharides that are considered biologically active. This study aimed to determine the feasibility of enhancing exopolysaccharide (EPS) production by ABSC in shake flask culture by supplementing the fermentation medium with anthocyanin extract. Different concentrations of Lycium ruthenicum Murr. (LRM) anthocyanin crude extract were tested on ABSC fermentation. The activity of phosphoglucose isomerase (PGI), phosphoglucose mutase (PGM), and phosphomannose isomerase (PMI), enzymes presumably involved in EPS synthesis by ABSC, was determined. ABSC transcriptomic profile in response to the presence of anthocyanins during fermentation was also investigated. LRM anthocyanin crude extract (0.06 mg/mL) was most effective in increasing EPS content and mycelial biomass (by 208.10% and 105.30%, respectively, P < 0.01). The activity of PGI, PGM, and PMI was increased in a medium where LRM anthocyanin extract and its main components (proanthocyanidins and petunia anthocyanin) were added. RNA-Seq analysis showed that 349 genes of ABSC were differentially expressed during fermentation in the medium containing anthocyanin extract of LRM; 93 genes were up-regulated and 256 genes down-regulated. From gene ontology enrichment analysis, differentially expressed genes were mostly assigned to carbohydrate metabolism and signal transduction categories. Collectively, LRM anthocyanins extract positively affected EPS production and mycelial biomass during ABSC fermentation. Our study provides a novel strategy for improving EPS production and mycelial growth during ABSC liquid submerged fermentation.

Secreted phosphoglucose isomerase is a novel biomarker of nonalcoholic fatty liver in mice and humans

The current gold standard for diagnosis of nonalcoholic fatty liver disease (NAFLD) and nonalcoholic steatohepatitis (NASH) is through a liver biopsy, and there is an urgent need to develop non-invasive methods for early detection. We previously demonstrated metabolic remodeling in the mouse fatty liver, which is marked by increased hepatic expression and activities of phosphoglucose isomerase (PGI) and several other glycolytic enzymes. Since PGI is actively transported out of the cell, acting as a multifunctional cytokine referred to as autocrine motility factor (AMF), we explored the possibility that PGI secreted from the fatty liver may be targeted for early detection of the silent disease. We report here that mice with NASH exhibited significantly elevated serum PGI enzyme activities compared to normal control (P < 0.005). We further confirmed the finding using serum/plasma samples (n = 73) collected from a cohort of NASH patients who were diagnosed according to Kleiner’s criteria, showing a normal mean PGI of 19.5 ± 8.8 IU/L and patient mean PGI of 105.6 ± 79.9 IU/L (P < 0.005). In addition, elevated blood PGI in NASH patients coincided with increased blood L-lactate. Cell culture experiments were then conducted to delineate the PGI-lactate axis, which revealed that treatment of HepG2 cells with recombinant PGI protein stimulated glycolysis and lactate output, suggesting that the disease-induced PGI likely contributed to the increased lactate in NASH patients. Taken together, the preclinical and clinical data validate secreted PGI as a useful biomarker of the fatty liver that can be easily screened at the point of care.

A Sulfoglycolytic Entner-Doudoroff Pathway in Rhizobium leguminosarum bv. trifolii SRDI565.

Rhizobia are nitrogen-fixing bacteria that engage in symbiotic relationships with plant hosts but can also persist as free-living bacteria in the soil and rhizosphere. Here, we show that free-living Rhizobium leguminosarum SRDI565 can grow on the sulfosugar sulfoquinovose (SQ) or the related glycoside SQ-glycerol using a sulfoglycolytic Entner-Doudoroff (sulfo-ED) pathway, resulting in production of sulfolactate (SL) as the major metabolic end product. Comparative proteomics supports the involvement of a sulfo-ED operon encoding an ABC transporter, sulfo-ED enzymes, and an SL exporter. Consistent with an oligotrophic lifestyle, proteomics data revealed little change in expression of the sulfo-ED proteins during growth on SQ versus mannitol, a result confirmed through biochemical assay of sulfoquinovosidase activity in cell lysates. Metabolomics analysis showed that growth on SQ involves gluconeogenesis to satisfy metabolic requirements for glucose-6-phosphate and fructose-6-phosphate. Metabolomics analysis also revealed the unexpected production of small amounts of sulfofructose and 2,3-dihydroxypropanesulfonate, which are proposed to arise from promiscuous activities of the glycolytic enzyme phosphoglucose isomerase and a nonspecific aldehyde reductase, respectively. The discovery of a rhizobium isolate with the ability to degrade SQ builds our knowledge of how these important symbiotic bacteria persist within soil.IMPORTANCE Sulfonate sulfur is a major form of organic sulfur in soils but requires biomineralization before it can be utilized by plants. Very little is known about the biochemical processes used to mobilize sulfonate sulfur. We show that a rhizobial isolate from soil, Rhizobium leguminosarum SRDI565, possesses the ability to degrade the abundant phototroph-derived carbohydrate sulfonate SQ through a sulfoglycolytic Entner-Doudoroff pathway. Proteomics and metabolomics demonstrated the utilization of this pathway during growth on SQ and provided evidence for gluconeogenesis. Unexpectedly, off-cycle sulfoglycolytic species were also detected, pointing to the complexity of metabolic processes within cells under conditions of sulfoglycolysis. Thus, rhizobial metabolism of the abundant sulfosugar SQ may contribute to persistence of the bacteria in the soil and to mobilization of sulfur in the pedosphere.

Glucose 6-Phosphate Accumulates via Phosphoglucose Isomerase Inhibition in Heart Muscle.

Metabolic and structural remodeling is a hallmark of heart failure. This remodeling involves activation of the mTOR (mammalian target of rapamycin) signaling pathway, but little is known on how intermediary metabolites are integrated as metabolic signals. We investigated the metabolic control of cardiac glycolysis and explored the potential of glucose 6-phosphate (G6P) to regulate glycolytic flux and mTOR activation. We developed a kinetic model of cardiomyocyte carbohydrate metabolism, CardioGlyco, to study the metabolic control of myocardial glycolysis and G6P levels. Metabolic control analysis revealed that G6P concentration is dependent on phosphoglucose isomerase (PGI) activity. Next, we integrated ex vivo tracer studies with mathematical simulations to test how changes in glucose supply and glycolytic flux affect mTOR activation. Nutrient deprivation promoted a tight coupling between glucose uptake and oxidation, G6P reduction, and increased protein-protein interaction between hexokinase II and mTOR. We validated the in silico modeling in cultured adult mouse ventricular cardiomyocytes by modulating PGI activity using erythrose 4-phosphate. Inhibition of glycolytic flux at the level of PGI caused G6P accumulation, which correlated with increased mTOR activation. Using click chemistry, we labeled newly synthesized proteins and confirmed that inhibition of PGI increases protein synthesis. The reduction of PGI activity directly affects myocyte growth by regulating mTOR activation.

Genome-Wide Association Mapping Reveals That Specific and Pleiotropic Regulatory Mechanisms Fine-Tune Central Metabolism and Growth in Arabidopsis.

Central metabolism is a coordinated network that is regulated at multiple levels by resource availability and by environmental and developmental cues. Its genetic architecture has been investigated by mapping metabolite quantitative trait loci (QTL). A more direct approach is to identify enzyme activity QTL, which distinguishes between cis-QTL in structural genes encoding enzymes and regulatory trans-QTL. Using genome-wide association studies, we mapped QTL for 24 enzyme activities, nine metabolites, three structural components, and biomass in Arabidopsis thaliana We detected strong cis-QTL for five enzyme activities. A cis-QTL for UDP-glucose pyrophosphorylase activity in the UGP1 promoter is maintained through balancing selection. Variation in acid invertase activity reflects multiple evolutionary events in the promoter and coding region of VAC-INVcis-QTL were also detected for ADP-glucose pyrophosphorylase, fumarase, and phosphoglucose isomerase activity. We detected many trans-QTL, including transcription factors, E3 ligases, protein targeting components, and protein kinases, and validated some by knockout analysis. trans-QTL are more frequent but tend to have smaller individual effects than cis-QTL. We detected many colocalized QTL, including a multitrait QTL on chromosome 4 that affects six enzyme activities, three metabolites, protein, and biomass. These traits are coordinately modified by different ACCELERATED CELL DEATH6 alleles, revealing a trade-off between metabolism and defense against biotic stress.

Carbon competition between fatty acids and starch during benzoin seeds maturation slows oil accumulation speed

The carbon flow balance is shifted to starch rather than oil accumulation during benzoin seed development stage (80–120 DAF) and thereby lipid deposition speed slows down. Benzoin [Styrax tonkinensis (Pierre) Craib ex Hartwich] is a woody oilseed species with potential value as a biofuel feedstock. However, the continuous drop in seed crude fat concentration during maturation had remained unexplained until it was investigated in this study. Seed filling began 60 days after flowering (DAF), when the hexose-to-sucrose ratio dropped, correlating with substantial morphological changes in kernels. Oil accumulated 20 days earlier than starch, as the concentrations of free fatty acid (50–70 DAF) and diacylglycerols (50–80 DAF) increased rapidly. Fatty acid synthesis slowed between 80 and 120 days, correlating with a sharp increase in the speed of starch deposition, while triacylglycerol accumulation and the profile of fatty acids remained stable. Phosphoglucose isomerase (PGI), pyruvate dehydrogenase complex (PDHC), and malate dehydrogenase (MDH) activities dropped progressively together with acetyl coenzyme carboxylase (ACCase) after 70 DAF before the reserve storage priority shifted from fatty acids to starch. PGI, PDHC, MDH, ACCase, and diglyceride acyltransferase (DAGT) activities rose again at 120 DAF, after which the speed of fatty acid accumulation increased, correlating with a 10-day lag phase in starch accumulation. High glucose 6-phosphate dehydrogenase (G6PDH) activity was observed during oil deposition. Soluble protein continued accumulating as protease activity decreased during benzoin seed development, while the total amylase activity rose twice between 30 and 70 DAF before the ADP-glucose pyrophosphorylase activity and starch content soared high simultaneously after 70 DAF. The collective results suggest that carbon partitioning between starch and lipid accumulation occurs during benzoin kernel development and is responsible for a reduction in the rate at which total fatty acids accumulate between 80 and 120 DAF.

Effects of mixed carbon sources on galactose and mannose content of exopolysaccharides and related enzyme activities in Ganoderma lucidum

This work investigated the effects of mixed carbon sources on the monosaccharide composition of exopolysaccharides (EPS) and activity of related enzymes, phosphoglucose isomerase (PGI), α-phosphoglucomutase (PGM), phosphomannose isomerase (PMI) and GDP-D-Man pyrophosphorylase (GMP), in the EPS biosynthesis by Ganoderma lucidum. Combinations of two carbon sources, glucose (Glc) + mannose (Man) and Glc + galactose (Gal), were applied at various mass ratios in the liquid culture of G. lucidum. The results showed that Glc, Gal, and Man were the major monosaccharides in G. lucidum EPS, and the combined mole percentages of monosaccharides were >80%, the same as those in the carbon source mixture. Activities of PGI, PMI, and GMP were correlated with Man mole percentage in EPSs and were enhanced by increasing the proportion of Gal for the carbon source Glc + Gal. PGM activity was correlated with Gal mole percentage and was reduced by increasing the Man proportion for Glc + Man. The expression of four genes encoding PGI, PMI, GMP and PGM reached peak levels on day 6 of culture, whereas the enzyme activities increased steadily from day 3 to 8. The monosaccharide composition of G. lucidum EPS for various mixed carbon source conditions thus appeared to be controlled by the translational level of genes encoding PGM, PGI, PMI, and GMP. These findings will be helpful to control the monosaccharide composition for desired biological activity of EPS produced by G. lucidum fermentation.

Influence of fermentation conditions on polysaccharide production and the activities of enzymes involved in the polysaccharide synthesis of Cordyceps militaris.

The influence of different fermentation conditions on intracellular polysaccharide (IPS) production and activities of the phosphoglucomutase (PGM), UDPG-pyrophosphorylase (UGP), phosphoglucose isomerase (PGI), UDPG-dehydrogenase (UGD), and glucokinase (GK) implicated in metabolite synthesis in Cordyceps militaris was evaluated. The highest IPS production (327.57 ± 6.27mg/100mL) was obtained when the strain was grown in the optimal medium containing glucose (40g · L(-1)), beef extract (10g · L(-1)), and CaCO3 (0.5g · L(-1)), and the initial pH and temperature were 7 and 25°C, respectively. The activities of PGM, UGP, and PGI were proved to be influenced by the fermentation conditions. A strong correlation between the activities of these enzymes and the production of IPS was found. The transcription level of the pgm gene (encoding PGM) was 1.049 times and 1.467 times compared to the ugp gene and pgi gene (encoding UGP and PGI), respectively, in the optimal culture medium. This result indicated that PGM might be the highly key enzyme to regulate the biosynthesis of IPS of C. militaris in a liquid-submerged culture. Our study might be helpful for further research on the pathway of polysaccharide biosynthesis aimed to improve the IPS production of C. militaris.

Sucrose fed-batch strategy enhanced biomass, polysaccharide, and ganoderic acids production in fermentation of Ganoderma lucidum 5.26.

Ganoderma, as a Chinese traditional medicine, has multiple bioactivities. However, industrial production was limited due to low yield during Ganoderma fermentation. In this work, sucrose was found to greatly enhance intracellular polysaccharide (IPS) content and specific extracellular polysaccharide (EPS) production rate. The mechanism was studied by analyzing the activities of enzymes related to polysaccharide biosynthesis. The results revealed that sucrose regulated the activities of phosphoglucomutase and phosphoglucose isomerase. When glucose and sucrose mixture was used as carbon source, biomass, polysaccharide and ganoderic acids (GAs) production was greatly enhanced. A sucrose fed-batch strategy was developed in 10-L bioreactor, and was scaled up to 300-L bioreactor. The biomass, EPS and IPS production was 25.5, 2.9 and 4.8 g/L, respectively, which was the highest biomass and IPS production in pilot scale. This study provides information for further understanding the regulation mechanism of Ganoderma polysaccharide biosynthesis. It demonstrates that sucrose fed-batch is a useful strategy for enhancing Ganoderma biomass, polysaccharide and GAs production.

AMF/PGI-mediated tumorigenesis through MAPK-ERK signaling in endometrial carcinoma.

Autocrine motility factor (AMF), which is also known as phosphoglucose isomerase (PGI), enhances tumor cell growth and motility. In this study, we found that AMF and its receptor were both highly expressed in Endometrial Carcinoma (EC) tissues compared to normal tissues. Levels of AMF were increased in serum of endometrial cancer patients. Downregulation of AMF by shRNA inhibited invasion, migration and proliferation as well as growth in a three-dimensional culture. AMF cytokine function, but not enzymatic activity of PGI, regulated tumorigenic activities of AMF. The MAPK-ERK1/2 pathway contributed to AMF-induced effects in EC cells. In agreement, Mek inhibitor decreased AMF-induced invasion, migration and proliferation of EC cells. In addition, in two mouse tumor metastasis models (EC cells delivered through left ventricle or intraperitoneally) AMF-silenced EC cells showed decreased tumor proliferative and metastatic capacities. We suggest that AMF/PGI is a potential therapeutic target in endometrial carcinoma.

Effects of culture conditions on monosaccharide composition of Ganoderma lucidum exopolysaccharide and on activities of related enzymes

We investigated the relationship between monosaccharide composition of Ganoderma lucidum exopolysaccharide (EPS) and activities of EPS synthesis enzymes under various culture temperatures and initial pH values. The mole percentages of three major EPS monosaccharides, glucose, galactose and mannose, varied depending on culture conditions and the resulting EPS displayed differing anti-tumor activities. In nine tested enzymes, higher enzyme activities were correlated with higher temperature and lower initial pH. Altered mole percentages of galactose and mannose under various culture conditions were associated with activities of α-phosphoglucomutase (PGM) and phosphoglucose isomerase (PGI), respectively, and that of mannose was also associated with phosphomannose isomerase (PMI) activity only under various pH. Our findings suggest that mole percentages of G. lucidum EPS monosaccharides can be manipulated by changes of culture conditions that affect enzyme activities, and that novel fermentation strategies based on this approach may enhance production and biological activity of EPS.

Impacts of drought stress on soluble carbohydrates and respiratory enzymes in fruit body of Auricularia auricula

In order to study the survival mechanisms to drought stress for fruit body of Auricularia auricula, soluble carbohydrates and respiratory enzymes were investigated. Fruit bodies were exposed to sunlight and were naturally dehydrated. Samples were taken at different levels of water loss (0%, 10%, 30%, 50% and 70%) to measure the content of soluble sugars and polysaccharides. The activities of phosphoglucose isomerase (PGI), combined glucose-6-phosphate dehydrogenase (G-6-PDH) and 6-phosphogluconate dehydrogenase (6-PGDH), and malate dehydrogenase (MDH), were also determined. The results showed that with the increase in water loss, soluble sugars and MDH activity declined, whereas the activities of G-6-PDH and 6-PGDH increased. Soluble polysaccharides content and PGI activity decreased with water loss up to 30% and increased afterwards. These results suggested that the pentose phosphate pathway (PPP), as demonstrated by activities of G-6-PDH and 6-PGDH, could be one of the mechanisms for survival during drought stress in the fruit body of A. auricula. Moreover, soluble polysaccharides may play a part in protecting the fruit body in further drought stress.

Rapid detection for the inhibition of phosphoglucose isomerase from Escherichia coli by mercury(II) chloride based on TLC-autographic analysis – preliminary studies

Abstract The quest for new techniques for screening inhibitors of phosphoglucose isomerase is crucially important owing to therapeutic control of chronic bacterial infections associated with the biosynthesis of bacterial biofilm. According to the new method, yellowish zones against the purple background could be visually observed where phosphoglucose isomerase activity was inhibited. The new protocol with NADPH/NBT/PMS staining for TLC-autographic method was able to detect PGI inhibition by pure reference substance as mercury(II) chloride.

Activities of fenbendazole in comparison with albendazole against Echinococcus multilocularis metacestodes in vitro and in a murine infection model

The current chemotherapeutic treatment of alveolar echinococcosis (AE) in humans is based on albendazole and/or mebendazole. However, the costs of treatment, life-long consumption of drugs, parasitostatic rather than parasiticidal activity of chemotherapy, and high recurrence rates after treatment interruption warrant more efficient treatment options. Experimental treatment of mice infected with Echinococcus multilocularis metacestodes with fenbendazole revealed similar efficacy to albendazole. Inspection of parasite tissue from infected and benzimidazole-treated mice by transmission electron microscopy (TEM) demonstrated drug-induced alterations within the germinal layer of the parasites, and most notably an almost complete absence of microtriches. On the other hand, upon in vitro exposure of metacestodes to benzimidazoles, no phosphoglucose isomerase activity could be detected in medium supernatants during treatment with any of these drugs, indicating that in vitro treatment did not severely affect the viability of metacestode tissue. Corresponding TEM analysis also revealed a dramatic shortening/retraction of microtriches as a hallmark of benzimidazole action, and as a consequence separation of the acellular laminated layer from the cellular germinal layer. Since TEM did not reveal any microtubule-based structures within Echinococcus microtriches, this effect cannot be explained by the previously described mechanism of action of benzimidazoles targeting β-tubulin, thus benzimidazoles must interact with additional targets that have not been yet identified. In addition, these results indicate the potential usefulness of fenbendazole for the chemotherapy of AE.

Effect of ultrafiltered fractions from casein on lactic acid biosynthesis and enzyme activity in yoghurt starter cultures

SummaryIn this work, five ultrafiltered fractions (UFs) with molecular mass less than 3 kDa (kilo‐daltons) from casein hydrolysates treated with alcalase, flavourzyme, neutrase, papain and trypsin, respectively, were obtained. The effect of five UFs on the fermentation for lactic acid (LA) production by mixed cultures of Streptococcus thermophilus (St) and Lactobacillus delbrueckii subsp. bulgaricus (Lb) during 72 h of cultivation was investigated. Compared with the control, LA production was, respectively, enhanced by 23.66%, 39.01%, 29.74%, 49.64% and 47.40% with the supplement of UF‐A, UF‐F, UF‐N, UF‐P and UF‐T at 24 h. The possible mechanism of LA production enhanced was elucidated by the time course analysis of the specific activity of glucokinase, phosphoglucose isomerase, 6‐phosphofructokinase, pyruvate kinase and lactate dehydrogenase during fermentation process. In addition, the results obtained showed the diverse influence of five UFs on the bacterial fermentation was attributed to their different amino acid distribution.