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- PDB-7yeh: Cryo-EM structure of human OGT-OGA complex -

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Basic information

Entry
Database: PDB / ID: 7yeh
TitleCryo-EM structure of human OGT-OGA complex
Components
  • Protein O-GlcNAcase
  • UDP-N-acetylglucosamine--peptide N-acetylglucosaminyltransferase 110 kDa subunit
KeywordsTRANSFERASE/HYDROLASE / O-GlcNAc transferase / O-GlcNAcase / Complex / Mutual inhibition / TRANSFERASE / TRANSFERASE-HYDROLASE complex
Function / homology
Function and homology information


glycoprotein metabolic process / protein N-acetylglucosaminyltransferase complex / hyalurononglucosaminidase activity / protein O-GlcNAc transferase / N-acetylglucosamine metabolic process / regulation of insulin receptor signaling pathway / protein O-acetylglucosaminyltransferase activity / positive regulation of transcription from RNA polymerase II promoter by glucose / glycoprotein catabolic process / protein O-GlcNAcase ...glycoprotein metabolic process / protein N-acetylglucosaminyltransferase complex / hyalurononglucosaminidase activity / protein O-GlcNAc transferase / N-acetylglucosamine metabolic process / regulation of insulin receptor signaling pathway / protein O-acetylglucosaminyltransferase activity / positive regulation of transcription from RNA polymerase II promoter by glucose / glycoprotein catabolic process / protein O-GlcNAcase / : / : / [protein]-3-O-(N-acetyl-D-glucosaminyl)-L-serine/L-threonine O-N-acetyl-alpha-D-glucosaminase activity / acetylglucosaminyltransferase activity / regulation of necroptotic process / regulation of Rac protein signal transduction / negative regulation of stem cell population maintenance / protein O-linked glycosylation / protein deglycosylation / NSL complex / : / regulation of neurotransmitter receptor localization to postsynaptic specialization membrane / regulation of glycolytic process / RIPK1-mediated regulated necrosis / regulation of synapse assembly / regulation of gluconeogenesis / positive regulation of stem cell population maintenance / Formation of WDR5-containing histone-modifying complexes / positive regulation of proteolysis / phosphatidylinositol-3,4,5-trisphosphate binding / mitophagy / hemopoiesis / negative regulation of proteasomal ubiquitin-dependent protein catabolic process / histone acetyltransferase complex / positive regulation of lipid biosynthetic process / negative regulation of protein ubiquitination / positive regulation of TORC1 signaling / response to nutrient / negative regulation of cell migration / positive regulation of translation / cell projection / beta-N-acetylglucosaminidase activity / mitochondrial membrane / cellular response to glucose stimulus / negative regulation of transforming growth factor beta receptor signaling pathway / circadian regulation of gene expression / response to insulin / Regulation of necroptotic cell death / protein processing / chromatin DNA binding / UCH proteinases / positive regulation of cold-induced thermogenesis / chromatin organization / HATs acetylate histones / glutamatergic synapse / apoptotic process / regulation of transcription by RNA polymerase II / positive regulation of DNA-templated transcription / negative regulation of transcription by RNA polymerase II / signal transduction / positive regulation of transcription by RNA polymerase II / protein-containing complex / nucleoplasm / membrane / identical protein binding / nucleus / plasma membrane / cytosol
Similarity search - Function
O-GlcNAc transferase, C-terminal / Glycosyl transferase family 41 / Glycosyl hydrolases family 84 (GH84) domain profile. / Beta-N-acetylglucosaminidase / beta-N-acetylglucosaminidase / TPR repeat / Tetratricopeptide repeat / Tetratricopeptide repeat 1 / Tetratricopeptide repeat / Tetratricopeptide repeat ...O-GlcNAc transferase, C-terminal / Glycosyl transferase family 41 / Glycosyl hydrolases family 84 (GH84) domain profile. / Beta-N-acetylglucosaminidase / beta-N-acetylglucosaminidase / TPR repeat / Tetratricopeptide repeat / Tetratricopeptide repeat 1 / Tetratricopeptide repeat / Tetratricopeptide repeat / TPR repeat region circular profile. / TPR repeat profile. / Acyl-CoA N-acyltransferase / Tetratricopeptide repeats / Tetratricopeptide repeat / Tetratricopeptide-like helical domain superfamily / Glycoside hydrolase superfamily
Similarity search - Domain/homology
URIDINE-5'-DIPHOSPHATE / UDP-N-acetylglucosamine--peptide N-acetylglucosaminyltransferase 110 kDa subunit / Protein O-GlcNAcase
Similarity search - Component
Biological speciesHomo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.92 Å
AuthorsLu, P. / Liu, Y. / Yu, H. / Gao, H.
Funding support China, 1items
OrganizationGrant numberCountry
National Natural Science Foundation of China (NSFC)32130053 China
CitationJournal: Nat Commun / Year: 2023
Title: Cryo-EM structure of human O-GlcNAcylation enzyme pair OGT-OGA complex.
Authors: Ping Lu / Yusong Liu / Maozhou He / Ting Cao / Mengquan Yang / Shutao Qi / Hongtao Yu / Haishan Gao /
Abstract: O-GlcNAcylation is a conserved post-translational modification that attaches N-acetyl glucosamine (GlcNAc) to myriad cellular proteins. In response to nutritional and hormonal signals, O- ...O-GlcNAcylation is a conserved post-translational modification that attaches N-acetyl glucosamine (GlcNAc) to myriad cellular proteins. In response to nutritional and hormonal signals, O-GlcNAcylation regulates diverse cellular processes by modulating the stability, structure, and function of target proteins. Dysregulation of O-GlcNAcylation has been implicated in the pathogenesis of cancer, diabetes, and neurodegeneration. A single pair of enzymes, the O-GlcNAc transferase (OGT) and O-GlcNAcase (OGA), catalyzes the addition and removal of O-GlcNAc on over 3,000 proteins in the human proteome. However, how OGT selects its native substrates and maintains the homeostatic control of O-GlcNAcylation of so many substrates against OGA is not fully understood. Here, we present the cryo-electron microscopy (cryo-EM) structures of human OGT and the OGT-OGA complex. Our studies reveal that OGT forms a functionally important scissor-shaped dimer. Within the OGT-OGA complex structure, a long flexible OGA segment occupies the extended substrate-binding groove of OGT and positions a serine for O-GlcNAcylation, thus preventing OGT from modifying other substrates. Conversely, OGT disrupts the functional dimerization of OGA and occludes its active site, resulting in the blocking of access by other substrates. This mutual inhibition between OGT and OGA may limit the futile O-GlcNAcylation cycles and help to maintain O-GlcNAc homeostasis.
History
DepositionJul 5, 2022Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0Jul 12, 2023Provider: repository / Type: Initial release
Revision 1.1Jan 24, 2024Group: Data collection / Database references / Refinement description
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / citation / citation_author / em_3d_fitting_list
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.journal_volume / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year / _em_3d_fitting_list.accession_code / _em_3d_fitting_list.initial_refinement_model_id / _em_3d_fitting_list.source_name / _em_3d_fitting_list.type

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
B: UDP-N-acetylglucosamine--peptide N-acetylglucosaminyltransferase 110 kDa subunit
A: UDP-N-acetylglucosamine--peptide N-acetylglucosaminyltransferase 110 kDa subunit
C: Protein O-GlcNAcase
D: Protein O-GlcNAcase
hetero molecules


Theoretical massNumber of molelcules
Total (without water)442,5746
Polymers441,9494
Non-polymers6252
Water0
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: gel filtration
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: Protein UDP-N-acetylglucosamine--peptide N-acetylglucosaminyltransferase 110 kDa subunit / O-GlcNAc transferase subunit p110 / O-linked N-acetylglucosamine transferase 110 kDa subunit / OGT


Mass: 117953.414 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: OGT
Production host: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria)
References: UniProt: O15294, protein O-GlcNAc transferase
#2: Protein Protein O-GlcNAcase / / OGA / Beta-N-acetylglucosaminidase / Beta-N-acetylhexosaminidase / Beta-hexosaminidase / Meningioma- ...OGA / Beta-N-acetylglucosaminidase / Beta-N-acetylhexosaminidase / Beta-hexosaminidase / Meningioma-expressed antigen 5 / N-acetyl-beta-D-glucosaminidase / N-acetyl-beta-glucosaminidase / Nuclear cytoplasmic O-GlcNAcase and acetyltransferase / NCOAT


Mass: 103020.906 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: OGA, HEXC, KIAA0679, MEA5, MGEA5
Production host: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria)
References: UniProt: O60502, protein O-GlcNAcase
#3: Chemical ChemComp-UDP / URIDINE-5'-DIPHOSPHATE / Uridine diphosphate


Type: RNA linking / Mass: 404.161 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C9H14N2O12P2 / Feature type: SUBJECT OF INVESTIGATION / Comment: UDP*YM
#4: Sugar ChemComp-NAG / 2-acetamido-2-deoxy-beta-D-glucopyranose / N-acetyl-beta-D-glucosamine / 2-acetamido-2-deoxy-beta-D-glucose / 2-acetamido-2-deoxy-D-glucose / 2-acetamido-2-deoxy-glucose / N-ACETYL-D-GLUCOSAMINE / N-Acetylglucosamine


Type: D-saccharide, beta linking / Mass: 221.208 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C8H15NO6 / Feature type: SUBJECT OF INVESTIGATION
IdentifierTypeProgram
DGlcpNAcbCONDENSED IUPAC CARBOHYDRATE SYMBOLGMML 1.0
N-acetyl-b-D-glucopyranosamineCOMMON NAMEGMML 1.0
b-D-GlcpNAcIUPAC CARBOHYDRATE SYMBOLPDB-CARE 1.0
GlcNAcSNFG CARBOHYDRATE SYMBOLGMML 1.0
Has ligand of interestY

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

ComponentName: Cryo-EM structure of human OGT-OGA complex / Type: COMPLEX / Entity ID: #1-#2 / Source: RECOMBINANT
Molecular weightValue: 0.4 MDa / Experimental value: NO
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Organism: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria)
Buffer solutionpH: 7.5
SpecimenConc.: 5 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: COPPER / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R1.2/1.3
VitrificationCryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277 K

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 2300 nm / Nominal defocus min: 1300 nm / Alignment procedure: COMA FREE
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingElectron dose: 50 e/Å2 / Film or detector model: GATAN K3 (6k x 4k)

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Processing

SoftwareName: PHENIX / Version: 1.18.2_3874: / Classification: refinement
EM software
IDNameCategory
4GctfCTF correction
7UCSF Chimeramodel fitting
9cryoSPARCinitial Euler assignment
10cryoSPARCfinal Euler assignment
11cryoSPARCclassification
12cryoSPARC3D reconstruction
13PHENIXmodel refinement
CTF correctionType: NONE
Particle selectionNum. of particles selected: 114543
SymmetryPoint symmetry: C1 (asymmetric)
3D reconstructionResolution: 3.92 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 114543 / Symmetry type: POINT
Atomic model buildingB value: 146 / Protocol: RIGID BODY FIT / Space: REAL
Atomic model building

3D fitting-ID: 1 / Pdb chain-ID: A / Source name: PDB / Type: experimental model

IDPDB-IDAccession codeInitial refinement model-ID
11W3B

1w3b

1W3B1
23PE3

3pe3

3PE32
35UN9

5un9

5UN93
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00619597
ELECTRON MICROSCOPYf_angle_d0.9626621
ELECTRON MICROSCOPYf_dihedral_angle_d11.12633
ELECTRON MICROSCOPYf_chiral_restr0.0492932
ELECTRON MICROSCOPYf_plane_restr0.0053470

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