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Food Chemistry: X Mar 2024The objective of this work was to investigate the influence of phosphoglycerate kinase-1 (PGK1) and pyruvate kinase-M2 (PKM2) activity on glycolysis, myofibrillar...
The objective of this work was to investigate the influence of phosphoglycerate kinase-1 (PGK1) and pyruvate kinase-M2 (PKM2) activity on glycolysis, myofibrillar proteins, calpain system, and apoptosis pathways of postmortem muscle. The activity of PGK1 and PKM2 was regulated by their inhibitors and activators to construct the postmortem glycolysis model and then incubated at 4 °C for 24 h. The results showed that compared to PGK1 and PKM2 inhibitors groups, the addition of PGK1 and PKM2 activators could accelerate glycogen consumption, ATP and lactate production, while declining pH value. Moreover, the addition of PGK1 and PKM2 activators could increase desmin degradation, μ-calpain activity, and caspase-3 abundance. Interestingly, troponin-T degradation was significantly increased both in PKM2 inhibitor and activator groups. It was suggested that PGK1 and PKM2 might be used as robust indicators to regulate meat quality by affecting the glycolysis, myofibrillar proteins, μ-calpain and apoptosis pathways in postmortem muscle.
PubMed: 38292674
DOI: 10.1016/j.fochx.2024.101125 -
Experimental Parasitology Apr 2018T. rangeli epimastigotes contain only a single detectable phosphoglycerate kinase (PGK) enzyme in their cytosol. Analysis of this parasite's recently sequenced genome...
T. rangeli epimastigotes contain only a single detectable phosphoglycerate kinase (PGK) enzyme in their cytosol. Analysis of this parasite's recently sequenced genome showed a gene predicted to code for a PGK with the same molecular mass as the natural enzyme, and with a cytosolic localization as well. In this work, we have partially purified the natural PGK from T. rangeli epimastigotes. Furthermore, we cloned the predicted PGK gene and expressed it as a recombinant active enzyme. Both purified enzymes were kinetically characterized and displayed similar substrate affinities, with Km values of 0.13 mM and 0.5 mM, and Km values of 0.28 mM and 0.71 mM, for the natural and recombinant enzyme, respectively. The optimal pH for activity of both enzymes was in the range of 8-10. Like other PGKs, TrPGK is monomeric with a molecular mass of approximately 44 kDa. The enzyme's kinetic characteristics are comparable with those of cytosolic PGK isoforms from related trypanosomatid species, indicating that, most likely, this enzyme is equivalent with the PGKB that is responsible for generating ATP in the cytosol of other trypanosomatids. This is the first report of a glycolytic enzyme characterization from T. rangeli.
Topics: Amino Acid Sequence; Base Sequence; Chromatography, Gel; Chromatography, Ion Exchange; Cloning, Molecular; Consensus Sequence; Cytosol; DNA, Intergenic; Hydrogen-Ion Concentration; Isoenzymes; Kinetics; Phosphoglycerate Kinase; Recombinant Proteins; Sequence Alignment; Trypanosoma rangeli
PubMed: 29526574
DOI: 10.1016/j.exppara.2018.03.009 -
Orphanet Journal of Rare Diseases Oct 2021Congenital hemolytic anemias (CHAs) comprise defects of the erythrocyte membrane proteins and of red blood cell enzymes metabolism, along with alterations of... (Review)
Review
Congenital hemolytic anemias (CHAs) comprise defects of the erythrocyte membrane proteins and of red blood cell enzymes metabolism, along with alterations of erythropoiesis. These rare and heterogeneous conditions may generate several difficulties from the diagnostic point of view. Membrane defects include hereditary spherocytosis and elliptocytosis, and the group of hereditary stomatocytosis; glucose-6-phosphate dehydrogenase and pyruvate kinase, are the most common enzyme deficiencies. Among ultra-rare forms, it is worth reminding other enzyme defects (glucosephosphate isomerase, phosphofructokinase, adenylate kinase, triosephosphate isomerase, phosphoglycerate kinase, hexokinase, and pyrimidine 5'-nucleotidase), and congenital dyserythropoietic anemias. Family history, clinical findings (anemia, hemolysis, splenomegaly, gallstones, and iron overload), red cells morphology, and biochemical tests are well recognized diagnostic tools. Molecular findings are increasingly used, particularly in recessive and de novo cases, and may be fundamental in unraveling the diagnosis. Notably, several confounders may further challenge the diagnostic workup, including concomitant blood loss, nutrients deficiency, alterations of hemolytic markers due to other causes (alloimmunization, infectious agents, rare metabolic disorders), coexistence of other hemolytic disorders (autoimmune hemolytic anemia, paroxysmal nocturnal hemoglobinuria, etc.). Additional factors to be considered are the possible association with bone marrow, renal or hepatic diseases, other causes of iron overload (hereditary hemochromatosis, hemoglobinopathies, metabolic diseases), and the presence of extra-hematological signs/symptoms. In this review we provide some instructive clinical vignettes that highlight the difficulties and confounders encountered in the diagnosis and clinical management of CHAs.
Topics: Anemia, Hemolytic, Congenital; Erythrocytes; Hemoglobinopathies; Humans; Pyruvate Kinase; Spherocytosis, Hereditary
PubMed: 34627331
DOI: 10.1186/s13023-021-02036-4 -
Current Protein & Peptide Science 2016Metabolic pathways that extract energy from carbon compounds are essential for an organism's survival. Therefore, inhibition of enzymes in these pathways represents a... (Review)
Review
Metabolic pathways that extract energy from carbon compounds are essential for an organism's survival. Therefore, inhibition of enzymes in these pathways represents a potential therapeutic strategy to combat parasitic infections. However, the high degree of similarity between host and parasite enzymes makes this strategy potentially difficult. Nevertheless, several existing drugs to treat infections by parasitic helminths (worms) target metabolic enzymes. These include the trivalent antimonials that target phosphofructokinase and Clorsulon that targets phosphoglycerate mutase and phosphoglycerate kinase. Glycolytic enzymes from a variety of helminths have been characterised biochemically, and some inhibitors identified. To date none of these inhibitors have been developed into therapies. Many of these enzymes are externalised from the parasite and so are also of interest in the development of potential vaccines. Less work has been done on tricarboxylic acid cycle enzymes and oxidative phosphorylation complexes. Again, while some inhibitors have been identified none have been developed into drug-like molecules. Barriers to the development of novel drugs targeting metabolic enzymes include the lack of experimentally determined structures of helminth enzymes, lack of direct proof that the enzymes are vital in the parasites and lack of cell culture systems for many helminth species. Nevertheless, the success of Clorsulon (which discriminates between highly similar host and parasite enzymes) should inspire us to consider making serious efforts to discover novel anthelminthics, which target metabolic enzymes.
Topics: Animals; Citric Acid Cycle; Drug Discovery; Glycolysis; Helminths; Humans; Molecular Targeted Therapy; Oxidative Phosphorylation
PubMed: 26983888
DOI: 10.2174/1389203717999160226180733 -
Biochimie Oct 2019Phosphoglycerate kinase (PGK) is a key enzyme of glycolysis which also acts as a mediator of DNA replication and repair in the nucleus. We have cloned and expressed PGK...
Phosphoglycerate kinase (PGK) is a key enzyme of glycolysis which also acts as a mediator of DNA replication and repair in the nucleus. We have cloned and expressed PGK in Brugia malayi. The rBmPGK was found to be 415 amino acid residues long having 45 kDa subunit molecular weight. This enzyme was also identified in different life stages of bovine filarial parasite Setaria cervi. The enzyme activity was highest in microfilarial stage followed by adult female and male as also shown by real time PCR in the present study. Further using BmPGK primers the cDNA prepared from S. cervi was amplified and sequenced which showed 100% homology with Brugia malayi PGK. B. malayi and S. cervi, PGK consists of conserved calmodulin binding domain (CaMBD) having 21 amino acids. In the present study we have shown the CaMBD binds to calcium-calmodulin and regulates its activity. The binding of calmodulin (CaM) with CaMBD was confirmed using calmodulin agarose binding pull down assay, which showed that the rBmPGK binds to CaM agarose-calcium dependent manner. The effect of CaM-Caon the activity of rBmPGK was studied at different concentration of CaM (0.01-5.0 μM) and calcium chloride (0.01-100 μM). The rBmPGK was activated up to 85% in the presence of CaM at 1 μM and 10 μM concentration of CaCl. Interestingly this activation was abrogated by metal chelator EDTA. Similar results were shown in case of Setaria cervi PGK. A significant increase (90 ± 10) % in ScPGK activity was observed in the presence of CaM and CaCl at 1.0 μM and 1.0 mM respectively, further increase in the conc. of CaCl, the activity of ScPGK was found to be decreased like rBmPGK. Bioinformatics studies have also confirmed the interaction between CaMBD and CaM which showed CaM interacted to Phe 206, Gln 220, Arg 223 and Asn 224 of rBmPGK CaM binding domain. On the basis of these findings, it has been suggested that the activity of filarial PGK could be regulated in cells by Ca-CaM depending upon the concentration of calcium. To the best of our knowledge this is first report in filarial parasite.
Topics: Animals; Brugia malayi; Calcium; Calmodulin; Cattle; Phosphoglycerate Kinase; Protein Binding; Protein Domains; Setaria Nematode
PubMed: 31446011
DOI: 10.1016/j.biochi.2019.08.012 -
Journal of Experimental & Clinical... Dec 2023Within the tumor immune microenvironment (TME), tumor-associated macrophages (TAMs) are crucial in modulating polarization states to influence cancer development through...
BACKGROUND
Within the tumor immune microenvironment (TME), tumor-associated macrophages (TAMs) are crucial in modulating polarization states to influence cancer development through metabolic reprogramming. While long non-coding RNAs (lncRNAs) have been shown to play a pivotal role in the progression of various cancers, the underlying mechanisms by which lncRNAs alter M2 polarization through macrophage metabolism remodeling remain unelucidated.
METHODS
RNA sequencing was used to screen for differentially expressed lncRNAs in TAMs and normal tissue-resident macrophages (NTRMs) isolated from pancreatic ductal adenocarcinoma (PDAC) tissues, whilst RT-qPCR and FISH were employed to detect the expression level of SNHG17. Moreover, a series of in vivo and in vitro experiments were conducted to assess the functions of SNHG17 from TAMs in the polarization and glycolysis of M2-like macrophages and in the proliferation and metastasis of pancreatic cancer cells (PCs). Furthermore, Western blotting, RNA pull-down, mass spectrometry, RIP, and dual-luciferase assays were utilized to explore the underlying mechanism through which SNHG17 induces pro-tumor macrophage formation.
RESULTS
SNHG17 was substantially enriched in TAMs and was positively correlated with a worse prognosis in PDAC. Meanwhile, functional assays determined that SNHG17 promoted the malignant progression of PCs by enhancing M2 macrophage polarization and anaerobic glycolysis. Mechanistically, SNHG17 could sponge miR-628-5p to release PGK1 mRNA and concurrently interact with the PGK1 protein, activating the pro-tumorigenic function of PGK1 by enhancing phosphorylation at the T168A site of PGK1 through ERK1/2 recruitment. Lastly, SNHG17 knockdown could reverse the polarization status of macrophages in PDAC.
CONCLUSIONS
The present study illustrated the essential role of SNHG17 and its molecular mechanism in TAMs derived from PDAC, indicating that SNHG17 might be a viable target for PDAC immunotherapy.
Topics: Humans; Phosphorylation; RNA, Long Noncoding; Anaerobiosis; Cell Line, Tumor; Cell Proliferation; Pancreatic Neoplasms; Carcinoma, Pancreatic Ductal; Macrophages; Glycolysis; MicroRNAs; Tumor Microenvironment; Phosphoglycerate Kinase
PubMed: 38098044
DOI: 10.1186/s13046-023-02890-z -
Molecular Nutrition & Food Research Dec 2022As the tremendous increases in consumption of animal-derived food, endogenous hydrocortisone migrating along the food chain to organism arouses extensive attention. This...
SCOPE
As the tremendous increases in consumption of animal-derived food, endogenous hydrocortisone migrating along the food chain to organism arouses extensive attention. This study aims to investigate the cumulative impacts of dietary hydrocortisone intake and mechanistic understanding on metabolism of lipid nutrients.
METHODS AND RESULTS
A total of 120 porcine muscles samples with different concentrations of hydrocortisone are collected at three time points. An operational food chain simulation framework is constructed and 175 lipid molecules are identified by UHPLC-Q-Orbitrap HRMS. Compared to the control group, 66 lipid molecules are significantly different, including 17 triglycerides and 31 glycerophospholipids. Integrated analyses of lipidomics and proteomics indicate that hydrocortisone promotes adipose triglyceride lipase and hormone sensitive lipase activity to precondition for triglycerides hydrolysis. Quantitative lipidomics analysis shows the presence of hydrocortisone decreases the concentration of docosahexaenoic acid (3.66 ± 0.15-3.09 ± 0.12 mg kg ) and eicosapentanoic acid (0.54 ± 0.09-0.48 ± 0.06 mg kg ). A noteworthy increase of most saturated triglycerides concentration with the prolonging of time is observed.
CONCLUSIONS
Hydrocortisone originating from animal-derived food induces glycerophospholipids degradation and triglycerides hydrolysis through promoting adipose triglyceride lipase, hormone sensitive lipase, and phosphoglycerate kinase activity and further intervenes lipid nutrients utilization.
Topics: Animals; Swine; Hydrocortisone; Adipose Tissue; Triglycerides; Nutrients; Glycerophospholipids; Eating; Lipid Metabolism
PubMed: 36261391
DOI: 10.1002/mnfr.202200487 -
Plastic and Reconstructive Surgery Jun 2023Aerobic glycolysis (the Warburg effect) may play an important role in keloid pathogenesis, which may be aggravated by the hypoxic microenvironment in keloids....
BACKGROUND
Aerobic glycolysis (the Warburg effect) may play an important role in keloid pathogenesis, which may be aggravated by the hypoxic microenvironment in keloids. Phosphoglycerate kinase 1 (PGK1), a key glycolytic enzyme, is essential for cellular aerobic glycolysis, but its role in keloid formation remains unknown. This study aimed to detect PGK1 expression in keloid tissue and investigate the effects of inhibiting PGK1 expression on keloid fibroblasts (KFbs) under hypoxia and normoxia.
METHODS
Normal skin and keloid samples were separated into two parts, one was used for immunohistochemistry, and one for primary cell culture. PGK1 tissue expression was detected by immunohistochemistry. Reverse-transcriptase polymerase chain reaction and Western blotting were used to detect PGK1, GLUT1, LDHA, and COL1 expression, and glucose uptake and lactate production were detected with a microplate reader. Cell proliferation and apoptosis were investigated with IncuCyte and flow cytometry. Cell migration and invasion were detected with Transwell assays. Glycolytic function was explored with the Seahorse XF96 system.
RESULTS
Immunohistochemistry showed PGK1 overexpression in keloid tissue compared with normal skin tissue ( P < 0.05). Consistently, PGK1 expression was significantly higher in KFbs than in normal skin fibroblasts (NFbs), and hypoxia stimulated PGK1 expression in KFbs and NFbs ( P < 0.05). PGK1 knockdown significantly inhibited KFb glycolysis, proliferation, migration, invasion, glucose consumption, and lactate production ( P < 0.05). Furthermore, GLUT1, LDHA, and COL1 expression was decreased in KFbs compared with NFbs ( P < 0.05). In addition, suppressing PGK1 may mediate the PI3K/AKT pathway to down-regulate GLUT1, LDHA, and COL1 expression ( P < 0.05).
CONCLUSIONS
These findings provide new evidence that suppressing PGK1, inhibiting glycolysis, reduces KFb proliferation, migration, invasion, and type I collagen expression. Targeting PGK1 to inhibit the Warburg effect may be a new therapeutic strategy for keloids.
CLINICAL RELEVANCE STATEMENT
This article may provide new suggestions into the pathogenesis and treatment of keloids.
CLINICAL QUESTION/LEVEL OF EVIDENCE
Therapeutic, V.
Topics: Humans; Keloid; Glucose Transporter Type 1; Phosphatidylinositol 3-Kinases; Glycolysis; Lactates; Hypoxia; Cell Proliferation; Fibroblasts; Phosphoglycerate Kinase
PubMed: 36728674
DOI: 10.1097/PRS.0000000000010137 -
Revue Neurologique Nov 2023According to recent findings, Phosphoglycerate Kinase 1 (pgk-1) enzyme is linked to Parkinson's disease (PD). Mutations in the PGK-1 gene lead to decreases in the pgk-1... (Review)
Review
Importance of glucose and its metabolism in neurodegenerative disorder, as well as the combination of multiple therapeutic strategies targeting α-synuclein and neuroprotection in the treatment of Parkinson's disease.
According to recent findings, Phosphoglycerate Kinase 1 (pgk-1) enzyme is linked to Parkinson's disease (PD). Mutations in the PGK-1 gene lead to decreases in the pgk-1 enzyme which causes an imbalance in the levels of energy demand and supply. An increase in glycolytic adenosine triphosphate (ATP) production would help alleviate energy deficiency and sustain the acute energetic need of neurons. Neurodegeneration is caused by an imbalance or reduction in ATP levels. Recent data suggest that medications that increase glycolysis and neuroprotection can be used to treat PD. The current study focuses on treatment options for disorders associated with the pgk-1 enzyme, GLP-1, and A receptor which can be utilized to treat PD. A combination of metformin and terazosin, exenatide and meclizine, istradefylline and salbutamol treatments may benefit parkinsonism. The review also looked at potential target-specific new techniques that might assist in satisfying unfulfilled requirements in the treatment of PD.
PubMed: 38040547
DOI: 10.1016/j.neurol.2023.08.011 -
Autophagy Nov 2022Macroautophagy/autophagy is a finely-regulated process in which cytoplasm encapsulated within transient organelles termed autophagosomes is delivered to lysosomes or...
Phosphatidic acid suppresses autophagy through competitive inhibition by binding GAPC (glyceraldehyde-3-phosphate dehydrogenase) and PGK (phosphoglycerate kinase) proteins.
Macroautophagy/autophagy is a finely-regulated process in which cytoplasm encapsulated within transient organelles termed autophagosomes is delivered to lysosomes or vacuoles for degradation. Phospholipids, particularly phosphatidic acid (PA) that functions as a second messenger, play crucial and differential roles in autophagosome formation; however, the underlying mechanism remains largely unknown. Here we demonstrated that PA inhibits autophagy through competitive inhibition of the formation of ATG3 (autophagy-related)-ATG8e and ATG6-VPS34 (vacuolar protein sorting 34) complexes. PA bound to GAPC (glyceraldehyde-3-phosphate dehydrogenase) or PGK (phosphoglycerate kinase) and promoted their interaction with ATG3 or ATG6, which further attenuated the interactions of ATG3-ATG8e or ATG6-VPS34, respectively. Structural and mutational analyses revealed the mechanism of PA binding with GAPCs and PGK3, and that GAPCs or ATG8e competitively interacted with ATG3, and PGK3 or VPS34 competitively interacted with ATG6, at the same binding interface. These results elucidate the molecular mechanism of how PA inhibits autophagy through binding GAPC or PGK3 proteins and expand the understanding of the functional mode of PA, demonstrating the importance of phospholipids in plant autophagy and providing a new perspective for autophagy regulation by phospholipids. ATG: autophagy-related; BiFC: bimolecular fluorescence complementation; co-IP: co-immunoprecipitation; Con A: concanamycin A; ER: endoplasmic reticulum; EZ: elongation zone; FRET-FLIM: fluorescence resonance energy transfer with fluorescence lifetime imaging microscopy; GAPDH: glyceraldehyde-3-phosphate dehydrogenase; GST: glutathione S-transferase; MDC: monodansylcadaverine; MZ: meristem zone; PA: phosphatidic acid; PAS: phagophore assembly site; PC: phosphatidylcholine; PE: phosphatidylethanolamine; PGK3: phosphoglycerate kinase; PtdIns3K: phosphatidylinositol 3-kinase; PLD: phospholipase D; TEM: transmission electron microscopy; TOR: target of rapamycin; VPS34: vacuolar protein sorting 34; WT: wild type; Y2H: yeast two-hybrid.
Topics: Autophagy; Autophagy-Related Proteins; Glyceraldehyde-3-Phosphate Dehydrogenases; Lysosomes; Phosphatidic Acids; Phosphoglycerate Kinase; Ubiquitin-Conjugating Enzymes
PubMed: 35289711
DOI: 10.1080/15548627.2022.2046449