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The FEBS Journal Oct 2022Regulation of glycogen metabolism is of vital importance in organisms of all three kingdoms of life. Although the pathways involved in glycogen synthesis and degradation...
Regulation of glycogen metabolism is of vital importance in organisms of all three kingdoms of life. Although the pathways involved in glycogen synthesis and degradation are well known, many regulatory aspects around the metabolism of this polysaccharide remain undeciphered. Here, we used the unicellular cyanobacterium Synechocystis as a model to investigate how glycogen metabolism is regulated in nitrogen-starved dormant cells, which entirely rely on glycogen catabolism to resume growth upon nitrogen repletion. We identified phosphoglucomutase 1 (PGM1) as a key regulatory point in glycogen metabolism, and post-translational modification as an essential mechanism for controlling its activity. We could show that PGM1 is phosphorylated ata residue in the regulatory latch domain (Ser 47) during nitrogen starvation, which inhibits its activity. Inactivation of PGM1 by phosphorylation at Ser 47 prevents premature degradation of the glycogen stores and appears to be essential for survival of Synechocystis in the dormant state. Remarkably, this regulatory mechanism seems to be evolutionary conserved in PGM1 enzymes, from bacteria to humans.
Topics: Glycogen; Humans; Nitrogen; Phosphoglucomutase; Phosphorylation; Synechocystis
PubMed: 35509259
DOI: 10.1111/febs.16471 -
Bioscience Reports Apr 2020Cells lining the uterus are responsible for storage and secretion of carbohydrates to support early embryonic development. Histotrophic secretions contain glycogen and...
Cells lining the uterus are responsible for storage and secretion of carbohydrates to support early embryonic development. Histotrophic secretions contain glycogen and glycolytic products such as lactate and pyruvate. Insufficient carbohydrate storage as glycogen has been correlated with infertility in women. While it is clear that changes in estrogen (17-β-estradiol (E2)) and progesterone (P4) in vivo affect the distribution of glucose in the uterine cells and secretions, the biochemical mechanism(s) by which they affect this crucial allocation is not well understood. Furthermore, in cultured uterine cells, neither E2 nor P4 affect glycogen storage without insulin present. We hypothesized that P4 and E2 alone affect the activity of glycolytic enzymes, glucose and glycolytic flux to increase glycogen storage (E2) and catabolism (P4) and increase pyruvate and lactate levels in culture. We measured the rate of glucose uptake and glycolysis in a mink immortalized epithelial cell line (GMMe) after 24-h exposure to 10 μM P4 and 10 nM E2 (pharmacologic levels) at 5 mM glucose and determined the kinetic parameters (Vmax, Km) of all enzymes. While the activities of many glycolytic enzymes in GMMe cells were shown to be decreased by E2 treatment, in contrast, glucose uptake, glycolytic flux and metabolites levels were not affected by the treatments. The cellular rationale for P4- and E2-induced decreases in the activity of enzymes may be to prime the system for other regulators such as insulin. In vivo, E2 and P4 may be necessary but not sufficient signals for uterine cycle carbohydrate allocation.
Topics: Animals; Cell Line; Enzyme Assays; Epithelial Cells; Estradiol; Estrous Cycle; Female; Glucose; Glucosephosphate Dehydrogenase; Glycogen; Glycolysis; Kinetics; Mink; Models, Animal; Phosphoglucomutase; Progesterone; Uterus
PubMed: 32239183
DOI: 10.1042/BSR20193512 -
Journal of Applied Microbiology Aug 2011To evaluate the role of α-phosphoglucomutase (α-Pgm) and phosphoglucose isomerase (Pgi) activities in growth rate, sugar-phosphates, UDP-sugars and lactate...
AIMS
To evaluate the role of α-phosphoglucomutase (α-Pgm) and phosphoglucose isomerase (Pgi) activities in growth rate, sugar-phosphates, UDP-sugars and lactate biosynthesis in Lactobacillus casei.
METHODS AND RESULTS
The pgm and pgi genes coding for α-Pgm and Pgi activities in L. casei BL23, respectively, were identified, cloned and shown to be functional by homologous overexpression. In MRS fermentation medium with glucose, overexpression of pgm gene in L. casei resulted in a growth rate reduced to 75% and glucose-6P levels reduced to 47%. By contrast, with lactose, the growth rate was raised to 119%. An increment of α-Pgm activity had no significant effect on UDP-sugar levels. Remarkably, Pgi overexpression in L. casei grown in lactose or galactose resulted in almost a double growth rate with respect to the control strain. The increased Pgi activity also resulted in glucose-6P levels reduced to 25 and 59% of control strain cultured in glucose and lactose, respectively, and the fructose-6P levels were increased to 128% on glucose. UDP-glucose and UDP-galactose levels were reduced to 66 and 55%, respectively, of control strain levels cultured in galactose. In addition, the lactate yield increased to 115% in the strain overproducing Pgi grown in galactose.
CONCLUSIONS
The physiological amount of α-Pgm and Pgi activities is limited for L. casei growth on lactose, and lactose and galactose, respectively, and that limitation was overcome by pgm and pgi gene overexpression. The increment of α-Pgm and Pgi activities, respectively, resulted in modified levels of sugar-phosphates, sugar-nucleotides and lactate showing the modulation capacity of the carbon fluxes in L. casei at the level of the glycolytic intermediate glucose-6P.
SIGNIFICANCE AND IMPACT OF THE STUDY
Knowledge of the role of key enzymes in metabolic fluxes at the branching point between anabolic and catabolic pathways would allow a rational design of engineering strategies in L. casei.
Topics: Bacterial Proteins; Culture Media; DNA, Bacterial; Fermentation; Galactose; Gene Expression Regulation, Bacterial; Glucose-6-Phosphate; Glucose-6-Phosphate Isomerase; Glycolysis; Industrial Microbiology; Lactic Acid; Lacticaseibacillus casei; Lactose; Phosphoglucomutase; Plasmids; Uridine Diphosphate
PubMed: 21605291
DOI: 10.1111/j.1365-2672.2011.05045.x -
Molecules (Basel, Switzerland) Dec 2022An anti-biofilm that can inhibit the matrix of biofilm formation is necessary to prevent recurrent and chronic infection. This study aimed to design compounds with a...
An anti-biofilm that can inhibit the matrix of biofilm formation is necessary to prevent recurrent and chronic infection. This study aimed to design compounds with a new mechanism through competitive inhibitory activity against phosphomannomutase/phosphoglucomutase (PMM/PGM), using in vitro assessment and a computational (in silico) approach. The active site of PMM/PGM was assessed through molecular redocking using L-tartaric acid as the native ligand and other small molecules, such as glucaric acid, D-sorbitol, and ascorbic acid. The docking program set the small molecules to the active site, showing a stable complex formation. Analysis of structural similarity, bioavailability, absorption, distribution, metabolism, excretion, and toxicity properties proved the potential application of ligands as an anti-biofilm. In vitro assessment with crystal violet showed that the ligands could reach up to 95.87% inhibition at different concentrations. The nitrocellulose membrane and scanning electron microscopic visualization showed that the untreated biofilm was denser than the ligand-treated biofilm.
Topics: Pseudomonas aeruginosa; Ligands; Phosphoglucomutase; Catalytic Domain; Biofilms; Anti-Bacterial Agents
PubMed: 36558064
DOI: 10.3390/molecules27248935 -
The Journal of Biological Chemistry Jun 2022Aspergillus fumigatus is the causative agent of invasive aspergillosis, an infection with mortality rates of up to 50%. The glucan-rich cell wall of A. fumigatus is a...
Aspergillus fumigatus is the causative agent of invasive aspergillosis, an infection with mortality rates of up to 50%. The glucan-rich cell wall of A. fumigatus is a protective structure that is absent from human cells and is a potential target for antifungal treatments. Glucan is synthesized from the donor uridine diphosphate glucose, with the conversion of glucose-6-phosphate to glucose-1-phosphate by the enzyme phosphoglucomutase (PGM) representing a key step in its biosynthesis. Here, we explore the possibility of selectively targeting A. fumigatus PGM (AfPGM) as an antifungal treatment strategy. Using a promoter replacement strategy, we constructed a conditional pgm mutant and revealed that pgm is required for A. fumigatus growth and cell wall integrity. In addition, using a fragment screen, we identified the thiol-reactive compound isothiazolone fragment of PGM as targeting a cysteine residue not conserved in the human ortholog. Furthermore, through scaffold exploration, we synthesized a para-aryl derivative (ISFP10) and demonstrated that it inhibits AfPGM with an IC of 2 μM and exhibits 50-fold selectivity over the human enzyme. Taken together, our data provide genetic validation of PGM as a therapeutic target and suggest new avenues for inhibiting AfPGM using covalent inhibitors that could serve as tools for chemical validation.
Topics: Antifungal Agents; Aspergillosis; Aspergillus fumigatus; Glucans; Humans; Phosphoglucomutase
PubMed: 35504355
DOI: 10.1016/j.jbc.2022.102003 -
Veterinary Research Jul 2020Lipooligosaccharides (LOSs) are virulence determinants of Glaesserella parasuis, a pathogen of the respiratory tract of pigs. We previously reported that disruption of...
Lipooligosaccharides (LOSs) are virulence determinants of Glaesserella parasuis, a pathogen of the respiratory tract of pigs. We previously reported that disruption of the galU or galE gene in G. parasuis results in increased sensitivity to porcine serum, indicating that the galactose catabolism pathway is required for polysaccharide formation in G. parasuis. Here, we evaluated the role of the HAPS_0849 gene in LOS synthesis. The G. parasuis SC096 HAPS_0849 mutant produced a highly truncated LOS molecule, although a small fraction of intact LOS was still observed, and this mutant was found to be more sensitive to serum than the parental strain. HAPS_0849 was overexpressed and purified for biochemical assays, and this protein exhibited phosphoglucomutase (PGM) activity. Heterologous expression of a pgm gene from Escherichia coli in the HAPS_0849 mutant led to restoration of the wild-type LOS glycoform, further demonstrating the PGM function of HAPS_0849 in G. parasuis. The autoagglutination and biofilm formation ability of this strain were also investigated. Disruption of HAPS_0849 led to an increased tendency to autoagglutinate and form more biofilms, and these enhanced phenotypes were observed in the absence of glucose. In addition, LOSs from HAPS_0849, galU and lgtB mutants had similar truncated glycoforms, while LOSs from the galE and lex-1 mutants exhibited another type of defective LOS pattern. These findings imply that HAPS_0849 may function upstream of GalU in the generation of glucose 1-phosphate. In conclusion, our results preliminarily described the functions of HAPS_0849 in G. parasuis, and this gene was partially required for LOS synthesis.
Topics: Bacterial Proteins; Escherichia coli; Gene Expression Regulation, Bacterial; Haemophilus parasuis; Lipopolysaccharides; Microorganisms, Genetically-Modified; Phosphoglucomutase
PubMed: 32736655
DOI: 10.1186/s13567-020-00822-9 -
Journal of Medical Genetics Jun 1983The polymorphism of PGM1 and Gc was studied by isoelectric focusing in pulmonary tuberculosis patients and controls. For the Gc system, the phenotypic frequencies did...
The polymorphism of PGM1 and Gc was studied by isoelectric focusing in pulmonary tuberculosis patients and controls. For the Gc system, the phenotypic frequencies did not differ significantly, but the PGM1 system showed a significant difference in the tuberculosis patients compared to controls. The data presented here show the potential of newly developed techniques to distinguish subjects with susceptibility to disease.
Topics: Carrier Proteins; Humans; Isoenzymes; Phosphoglucomutase; Tuberculosis; Vitamin D-Binding Protein
PubMed: 6224013
DOI: 10.1136/jmg.20.3.220 -
Annals of Botany Nov 2023Crassulacean acid metabolism (CAM) is a specialized type of photosynthesis characterized by a diel pattern of stomatal opening at night and closure during the day, which...
The starch-deficient plastidic PHOSPHOGLUCOMUTASE mutant of the constitutive crassulacean acid metabolism (CAM) species Kalanchoë fedtschenkoi impacts diel regulation and timing of stomatal CO2 responsiveness.
BACKGROUND AND AIMS
Crassulacean acid metabolism (CAM) is a specialized type of photosynthesis characterized by a diel pattern of stomatal opening at night and closure during the day, which increases water-use efficiency. Starch degradation is a key regulator of CAM, providing phosphoenolpyruvate as a substrate in the mesophyll for nocturnal assimilation of CO2. Growing recognition of a key role for starch degradation in C3 photosynthesis guard cells for mediating daytime stomatal opening presents the possibility that starch degradation might also impact CAM by regulating the provision of energy and osmolytes to increase guard cell turgor and drive stomatal opening at night. In this study, we tested the hypothesis that the timing of diel starch turnover in CAM guard cells has been reprogrammed during evolution to enable nocturnal stomatal opening and daytime closure.
METHODS
Biochemical and genetic characterization of wild-type and starch-deficient RNAi lines of Kalanchoë fedtschenkoi with reduced activity of plastidic phosphoglucomutase (PGM) constituted a preliminary approach for the understanding of starch metabolism and its implications for stomatal regulation in CAM plants.
KEY RESULTS
Starch deficiency reduced nocturnal net CO2 uptake but had negligible impact on nocturnal stomatal opening. In contrast, daytime stomatal closure was reduced in magnitude and duration in the starch-deficient rPGM RNAi lines, and their stomata were unable to remain closed in response to elevated concentrations of atmospheric CO2 administered during the day. Curtailed daytime stomatal closure was linked to higher soluble sugar contents in the epidermis and mesophyll.
CONCLUSIONS
Nocturnal stomatal opening is not reliant upon starch degradation, but starch biosynthesis is an important sink for carbohydrates, ensuring daytime stomatal closure in this CAM species.
Topics: Crassulacean Acid Metabolism; Kalanchoe; Phosphoglucomutase; Carbon Dioxide; Starch; Photosynthesis
PubMed: 36661206
DOI: 10.1093/aob/mcad017 -
The Journal of Allergy and Clinical... May 2014Recurrent bacterial and fungal infections, eczema, and increased serum IgE levels characterize patients with the hyper-IgE syndrome (HIES). Known genetic causes for HIES... (Clinical Trial)
Clinical Trial
BACKGROUND
Recurrent bacterial and fungal infections, eczema, and increased serum IgE levels characterize patients with the hyper-IgE syndrome (HIES). Known genetic causes for HIES are mutations in signal transducer and activator of transcription 3 (STAT3) and dedicator of cytokinesis 8 (DOCK8), which are involved in signal transduction pathways. However, glycosylation defects have not been described in patients with HIES. One crucial enzyme in the glycosylation pathway is phosphoglucomutase 3 (PGM3), which catalyzes a key step in the synthesis of uridine diphosphate N-acetylglucosamine, which is required for the biosynthesis of N-glycans.
OBJECTIVE
We sought to elucidate the genetic cause in patients with HIES who do not carry mutations in STAT3 or DOCK8.
METHODS
After establishing a linkage interval by means of SNPchip genotyping and homozygosity mapping in 2 families with HIES from Tunisia, mutational analysis was performed with selector-based, high-throughput sequencing. Protein expression was analyzed by means of Western blotting, and glycosylation was profiled by using mass spectrometry.
RESULTS
Mutational analysis of candidate genes in an 11.9-Mb linkage region on chromosome 6 shared by 2 multiplex families identified 2 homozygous mutations in PGM3 that segregated with disease status and followed recessive inheritance. The mutations predict amino acid changes in PGM3 (p.Glu340del and p.Leu83Ser). A third homozygous mutation (p.Asp502Tyr) and the p.Leu83Ser variant were identified in 2 other affected families, respectively. These hypomorphic mutations have an effect on the biosynthetic reactions involving uridine diphosphate N-acetylglucosamine. Glycomic analysis revealed an aberrant glycosylation pattern in leukocytes demonstrated by a reduced level of tri-antennary and tetra-antennary N-glycans. T-cell proliferation and differentiation were impaired in patients. Most patients had developmental delay, and many had psychomotor retardation.
CONCLUSION
Impairment of PGM3 function leads to a novel primary (inborn) error of development and immunity because biallelic hypomorphic mutations are associated with impaired glycosylation and a hyper-IgE-like phenotype.
Topics: Adult; Amino Acid Substitution; Cell Proliferation; Child; Chromosomes, Human, Pair 6; Female; Genetic Diseases, Inborn; Genetic Linkage; Glycosylation; Homozygote; Humans; Immunity; Immunoglobulin E; Infant; Job Syndrome; Male; Mutation, Missense; Phosphoglucomutase; T-Lymphocytes; Tunisia
PubMed: 24698316
DOI: 10.1016/j.jaci.2014.02.025 -
Plant Physiology Jul 1998Phosphoglucomutase (PGM) catalyzes the interconversion of glucose (Glc)-1- and Glc-6-phosphate in the synthesis and consumption of sucrose. We isolated two maize (Zea...
Phosphoglucomutase (PGM) catalyzes the interconversion of glucose (Glc)-1- and Glc-6-phosphate in the synthesis and consumption of sucrose. We isolated two maize (Zea mays L.) cDNAs that encode PGM with 98.5% identity in their deduced amino acid sequence. Southern-blot analysis with genomic DNA from lines with different Pgm1 and Pgm2 genotypes suggested that the cDNAs encode the two known cytosolic PGM isozymes, PGM1 and PGM2. The cytosolic PGMs of maize are distinct from a plastidic PGM of spinach (Spinacia oleracea). The deduced amino acid sequences of the cytosolic PGMs contain the conserved phosphate-transfer catalytic center and the metal-ion-binding site of known prokaryotic and eukaryotic PGMs. PGM mRNA was detectable by RNA-blot analysis in all tissues and organs examined except silk. A reduction in PGM mRNA accumulation was detected in roots deprived of O2 for 24 h, along with reduced synthesis of a PGM identified as a 67-kD phosphoprotein on two-dimensional gels. Therefore, PGM is not one of the so-called "anaerobic polypeptides." Nevertheless, the specific activity of PGM was not significantly affected in roots deprived of O2 for 24 h. We propose that PGM is a stable protein and that existing levels are sufficient to maintain the flux of Glc-1-phosphate into glycolysis under O2 deprivation.
Topics: Aerobiosis; Amino Acid Sequence; Cloning, Molecular; Cytosol; Gene Expression Regulation, Developmental; Gene Expression Regulation, Enzymologic; Gene Expression Regulation, Plant; Genes, Plant; Hypoxia; Isoenzymes; Molecular Sequence Data; Multigene Family; Phosphoglucomutase; Phosphoproteins; Plant Roots; RNA, Messenger; Sequence Homology, Amino Acid; Transcription, Genetic; Zea mays
PubMed: 9662542
DOI: 10.1104/pp.117.3.997