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Autophagy Jul 2023Degradation of macromolecules delivered to lysosomes by processes such as autophagy or endocytosis is crucial for cellular function. Lysosomes require more than 60...
Degradation of macromolecules delivered to lysosomes by processes such as autophagy or endocytosis is crucial for cellular function. Lysosomes require more than 60 soluble hydrolases in order to catabolize such macromolecules. These soluble hydrolases are tagged with mannose6-phosphate (M6P) moieties in sequential reactions by the Golgi-resident GlcNAc-1-phosphotransferase complex and NAGPA/UCE/uncovering enzyme (N-acetylglucosamine-1-phosphodiester alpha-N-acetylglucosaminidase), which allows their delivery to endosomal/lysosomal compartments through trafficking mediated by cation-dependent and -independent mannose 6-phosphate receptors (MPRs). We and others recently identified TMEM251 as a novel regulator of the M6P pathway via independent genome-wide genetic screening strategies. We renamed TMEM251 to LYSET (lysosomal enzyme trafficking factor) to establish nomenclature reflective to this gene's function. LYSET is a Golgi-localized transmembrane protein important for the retention of the GlcNAc-1-phosphotransferase complex in the Golgi-apparatus. The current understanding of LYSET's importance regarding human biology is 3-fold: 1) highly pathogenic viruses that depend on lysosomal hydrolase activity require LYSET for infection. 2) The presence of LYSET is critical for cancer cell proliferation in nutrient-deprived environments in which extracellular proteins must be catabolized. 3) Inherited pathogenic alleles of LYSET can cause a severe inherited disease which resembles GlcNAc-1-phosphotransferase deficiency (i.e., mucolipidosis type II). GlcNAc-1-PT: GlcNAc-1-phosphotransferase; KO: knockout; LSD: lysosomal storage disorder; LYSET: lysosomal enzyme trafficking factor; M6P: mannose 6-phosphate; MPRs: mannose-6-phosphate receptors, cation-dependent or -independent; MBTPS1/site-1 protease: membrane bound transcription factor peptidase, site 1; MLII: mucolipidosis type II; WT: wild-type.
Topics: Humans; Mucolipidoses; Mannose; Autophagy; Lysosomes; Hydrolases; Receptor, IGF Type 2; Cations; Phosphotransferases
PubMed: 36633450
DOI: 10.1080/15548627.2023.2167376 -
Current Topics in Medicinal Chemistry 2020
Topics: Acetylcholinesterase; Butyrylcholinesterase; DNA Topoisomerases, Type II; Drug Design; Enzyme Inhibitors; Humans; Molecular Docking Simulation; Neuraminidase; Phosphotransferases
PubMed: 33054702
DOI: 10.2174/156802662023200917115547 -
Journal of Drug Targeting Jul 2021Breast cancer is the most common cancer in women. Despite advances in screening women for genetic predisposition to breast cancer and risk stratification, a majority of...
Breast cancer is the most common cancer in women. Despite advances in screening women for genetic predisposition to breast cancer and risk stratification, a majority of women carriers remain undetected until they become affected. Thus, there is a need to develop a cost-effective, rapid, sensitive and non-invasive early-stage diagnostic method. Kinases are involved in all fundamental cellular processes and mutations in kinases have been reported as drivers of cancer. PPARγ is a ligand-activated transcription factor that plays important roles in cell proliferation and metabolism. However, the complete set of kinases modulated by PPARγ is still unknown. In this study, we identified human kinases that are potential PPARγ targets and evaluated their differential expression and gene pair correlations in human breast cancer patient dataset TCGA-BRCA. We further confirmed the findings in human breast cancer cell lines MCF7 and SK-BR-3 using a kinome array. We observed that gene pair correlations are lost in tumours as compared to healthy controls and could be used as a supplement strategy for diagnosis and prognosis of breast cancer.
Topics: Breast Neoplasms; Cell Line, Tumor; Datasets as Topic; Female; Humans; MCF-7 Cells; Mutation; PPAR gamma; Phosphotransferases; Prognosis
PubMed: 33496213
DOI: 10.1080/1061186X.2021.1877719 -
Biochimica Et Biophysica Acta.... Nov 2020Mannose transporters constitute a superfamily (Man-PTS) of the Phosphoenolpyruvate Carbohydrate Phosphotransferase System (PTS). The membrane complexes are homotrimers... (Review)
Review
Mannose transporters constitute a superfamily (Man-PTS) of the Phosphoenolpyruvate Carbohydrate Phosphotransferase System (PTS). The membrane complexes are homotrimers of protomers consisting of two subunits, IIC and IID. The two subunits without recognizable sequence similarity assume the same fold, and in the protomer are structurally related by a two fold pseudosymmetry axis parallel to membrane-plane (Liu et al. (2019) Cell Research 29 680). Two reentrant loops and two transmembrane helices of each subunit together form the N-terminal transport domain. Two three-helix bundles, one of each subunit, form the scaffold domain. The protomer is stabilized by a helix swap between these bundles. The two C-terminal helices of IIC mediate the interprotomer contacts. PTS occur in bacteria and archaea but not in eukaryotes. Man-PTS are abundant in Gram-positive bacteria living on carbohydrate rich mucosal surfaces. A subgroup of IICIID complexes serve as receptors for class IIa bacteriocins and as channel for the penetration of bacteriophage lambda DNA across the inner membrane. Some Man-PTS are associated with host-pathogen and -symbiont processes.
Topics: Bacterial Proteins; Bacteriocins; Bacteriophages; Gram-Positive Bacteria; Mannose; Phosphotransferases; Protein Conformation, alpha-Helical; Protein Domains
PubMed: 32710850
DOI: 10.1016/j.bbamem.2020.183412 -
Journal of Agricultural and Food... Oct 2022Lactic acid bacteria are indispensable functional microorganisms for cereal vinegar brewing, but cell activities are inhibited by the dominant acetic acid stress....
Lactic acid bacteria are indispensable functional microorganisms for cereal vinegar brewing, but cell activities are inhibited by the dominant acetic acid stress. Herein, an acetic-acid-tolerant strain isolated previously was identified as , which also exhibited good resistance to other stresses during vinegar brewing. Proteomics analysis evidenced that differentially expressed proteins involved in the glycolysis and gluconeogenesis pathway, pyruvate metabolism, and sugar phosphotransferase system were all downregulated. Meanwhile, saturation of fatty acids and antioxidant enzymes was strengthened. The effects of several proteins on the resistance of and relied on the types of strain and stress. AccA and AcpP participating in fatty acid metabolism and biosynthesis and Mnc related to stress response were found to protect cells by modifying fatty acid compositions and reinforcing the antioxidant defense system. Our works deepen the mechanisms of under acetic acid and offer targets for engineering cell tolerance.
Topics: Acetic Acid; Antioxidants; Fermentation; Pediococcus; Pediococcus acidilactici; Phosphotransferases; Proteomics; Pyruvates; Sugars
PubMed: 36125361
DOI: 10.1021/acs.jafc.2c04160 -
Structure (London, England : 1993) May 2017The macrolides are a class of antibiotic, characterized by a large macrocyclic lactone ring that can be inactivated by macrolide phosphotransferase enzymes. We present...
The macrolides are a class of antibiotic, characterized by a large macrocyclic lactone ring that can be inactivated by macrolide phosphotransferase enzymes. We present structures for MPH(2')-I and MPH(2')-II in the apo state, and in complex with GTP analogs and six different macrolides. These represent the first structures from the two main classes of macrolide phosphotransferases. The structures show that the enzymes are related to the aminoglycoside phosphotransferases, but are distinguished from them by the presence of a large interdomain linker that contributes to an expanded antibiotic binding pocket. This pocket is largely hydrophobic, with a negatively charged patch located at a conserved aspartate residue, rationalizing the broad-spectrum resistance conferred by the enzymes. Complementary mutation studies provide insights into factors governing substrate specificity. A comparison with macrolides bound to their natural target, the 50S ribosome, suggests avenues for next-generation antibiotic development.
Topics: Bacteria; Bacterial Proteins; Binding Sites; Drug Resistance, Bacterial; Guanosine Triphosphate; Macrolides; Phosphotransferases (Alcohol Group Acceptor); Protein Binding; Substrate Specificity
PubMed: 28416110
DOI: 10.1016/j.str.2017.03.007 -
Current Opinion in Chemical Biology Dec 2021Advancements in chemical proteomics and mass spectrometry lipidomics are providing new opportunities to understand lipid kinase activity, specificity, and regulation on... (Review)
Review
Advancements in chemical proteomics and mass spectrometry lipidomics are providing new opportunities to understand lipid kinase activity, specificity, and regulation on a global cellular scale. Here, we describe recent developments in chemical biology of lipid kinases with a focus on those members that phosphorylate diacylglycerols. We further discuss future implications of how these mass spectrometry-based approaches can be adapted for studies of additional lipid kinase members with the aim of bridging the gap between protein and lipid kinase-focused investigations.
Topics: Diacylglycerol Kinase; Lipids; Mass Spectrometry; Phosphotransferases; Proteomics
PubMed: 34311404
DOI: 10.1016/j.cbpa.2021.06.007 -
Functional characterization of the phosphotransferase system in Parageobacillus thermoglucosidasius.Scientific Reports May 2023Parageobacillus thermoglucosidasius is a thermophilic bacterium characterized by rapid growth, low nutrient requirements, and amenability to genetic manipulation. These...
Parageobacillus thermoglucosidasius is a thermophilic bacterium characterized by rapid growth, low nutrient requirements, and amenability to genetic manipulation. These characteristics along with its ability to ferment a broad range of carbohydrates make P. thermoglucosidasius a potential workhorse in whole-cell biocatalysis. The phosphoenolpyruvate:carbohydrate phosphotransferase system (PTS) catalyzes the transport and phosphorylation of carbohydrates and sugar derivatives in bacteria, making it important for their physiological characterization. In this study, the role of PTS elements on the catabolism of PTS and non-PTS substrates was investigated for P. thermoglucosidasius DSM 2542. Knockout of the common enzyme I, part of all PTSs, showed that arbutin, cellobiose, fructose, glucose, glycerol, mannitol, mannose, N-acetylglucosamine, N-acetylmuramic acid, sorbitol, salicin, sucrose, and trehalose were PTS-dependent on translocation and coupled to phosphorylation. The role of each putative PTS was investigated and six PTS-deletion variants could not grow on arbutin, mannitol, N-acetylglucosamine, sorbitol, and trehalose as the main carbon source, or showed diminished growth on N-acetylmuramic acid. We concluded that PTS is a pivotal factor in the sugar metabolism of P. thermoglucosidasius and established six PTS variants important for the translocation of specific carbohydrates. This study lays the groundwork for engineering efforts with P. thermoglucosidasius towards efficient utilization of diverse carbon substrates for whole-cell biocatalysis.
Topics: Acetylglucosamine; Arbutin; Trehalose; Phosphotransferases; Carbohydrates; Bacteria; Mannitol; Sorbitol; Phosphoenolpyruvate Sugar Phosphotransferase System; Bacterial Proteins
PubMed: 37130962
DOI: 10.1038/s41598-023-33918-1 -
Topics in Current Chemistry (Cham) Apr 2017The phosphoryl group, PO, is the dynamic structural unit in the biological chemistry of phosphorus. Its transfer from a donor to an acceptor atom, with oxygen much more... (Review)
Review
The phosphoryl group, PO, is the dynamic structural unit in the biological chemistry of phosphorus. Its transfer from a donor to an acceptor atom, with oxygen much more prevalent than nitrogen, carbon, or sulfur, is at the core of a great majority of enzyme-catalyzed reactions involving phosphate esters, anhydrides, amidates, and phosphorothioates. The serendipitous discovery that the phosphoryl group could be labeled by "nuclear mutation," by substitution of PO by MgF or AlF, has underpinned the application of metal fluoride (MF ) complexes to mimic transition states for enzymatic phosphoryl transfer reactions, with sufficient stability for experimental analysis. Protein crystallography in the solid state and F NMR in solution have enabled direct observation of ternary and quaternary protein complexes embracing MF transition state models with precision. These studies have underpinned a radically new mechanistic approach to enzyme catalysis for a huge range of phosphoryl transfer processes, as varied as kinases, phosphatases, phosphomutases, and phosphohydrolases. The results, without exception, have endorsed trigonal bipyramidal geometry (tbp) for concerted, "in-line" stereochemistry of phosphoryl transfer. QM computations have established the validity of tbp MF complexes as reliable models for true transition states, delivering similar bond lengths, coordination to essential metal ions, and virtually identical hydrogen bond networks. The emergence of protein control of reactant orbital overlap between bond-forming species within enzyme transition states is a new challenging theme for wider exploration.
Topics: Aluminum Compounds; Fluorides; Magnesium Compounds; Molecular Structure; Phosphines; Phosphoric Monoester Hydrolases; Phosphotransferases; Phosphotransferases (Phosphomutases)
PubMed: 28299727
DOI: 10.1007/s41061-017-0130-y -
The Journal of Biological Chemistry Mar 2022GlcNAc-1-phosphotransferase catalyzes the initial step in the formation of the mannose-6-phosphate tag that labels ∼60 lysosomal proteins for transport. Mutations in...
GlcNAc-1-phosphotransferase catalyzes the initial step in the formation of the mannose-6-phosphate tag that labels ∼60 lysosomal proteins for transport. Mutations in GlcNAc-1-phosphotransferase are known to cause lysosomal storage disorders such as mucolipidoses. However, the molecular mechanism of GlcNAc-1-phosphotransferase activity remains unclear. Mammalian GlcNAc-1-phosphotransferases are α2β2γ2 hexamers in which the core catalytic α- and β-subunits are derived from the GNPTAB (N-acetylglucosamine-1-phosphate transferase subunits alpha and beta) gene. Here, we present the cryo-electron microscopy structure of the Drosophila melanogaster GNPTAB homolog, DmGNPTAB. We identified four conserved regions located far apart in the sequence that fold into the catalytic domain, which exhibits structural similarity to that of the UDP-glucose glycoprotein glucosyltransferase. Comparison with UDP-glucose glycoprotein glucosyltransferase also revealed a putative donor substrate-binding site, and the functional requirements of critical residues in human GNPTAB were validated using GNPTAB-knockout cells. Finally, we show that DmGNPTAB forms a homodimer that is evolutionarily conserved and that perturbing the dimer interface undermines the maturation and activity of human GNPTAB. These results provide important insights into GlcNAc-1-phosphotransferase function and related diseases.
Topics: Animals; Cryoelectron Microscopy; Drosophila melanogaster; Lysosomes; Mammals; Mucolipidoses; Proteins; Structure-Activity Relationship; Transferases (Other Substituted Phosphate Groups)
PubMed: 35148990
DOI: 10.1016/j.jbc.2022.101702