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- EMDB-38257: Cryo-EM structure of glutamate dehydrogenase from thermococcus pr... -
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Open data
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Basic information
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Title | Cryo-EM structure of glutamate dehydrogenase from thermococcus profundus incorporating NADPH and AKG in the steady stage of reaction | |||||||||||||||||||||||||||||||||||||||||||||
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![]() | Complex / Coenzyme / NADPH / 2-oxoglutarate / OXIDOREDUCTASE | |||||||||||||||||||||||||||||||||||||||||||||
Function / homology | ![]() glutamate dehydrogenase [NAD(P)+] / glutamate dehydrogenase (NADP+) activity / glutamate dehydrogenase (NAD+) activity / glutamate catabolic process Similarity search - Function | |||||||||||||||||||||||||||||||||||||||||||||
Biological species | ![]() ![]() | |||||||||||||||||||||||||||||||||||||||||||||
Method | single particle reconstruction / cryo EM / Resolution: 2.87 Å | |||||||||||||||||||||||||||||||||||||||||||||
![]() | Wakabayashi T / Oide M / Nakasako M | |||||||||||||||||||||||||||||||||||||||||||||
Funding support | ![]()
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![]() | ![]() Title: CryoEM-sampling of metastable conformations appearing in cofactor-ligand association and catalysis of glutamate dehydrogenase. Authors: Taiki Wakabayashi / Mao Oide / Masayoshi Nakasako / ![]() Abstract: Kinetic aspects of enzymatic reactions are described by equations based on the Michaelis-Menten theory for the initial stage. However, the kinetic parameters provide little information on the atomic ...Kinetic aspects of enzymatic reactions are described by equations based on the Michaelis-Menten theory for the initial stage. However, the kinetic parameters provide little information on the atomic mechanism of the reaction. In this study, we analyzed structures of glutamate dehydrogenase in the initial and steady stages of the reaction using cryoEM at near-atomic resolution. In the initial stage, four metastable conformations displayed different domain motions and cofactor/ligand association modes. The most striking finding was that the enzyme-cofactor-substrate complex, treated as a single state in the enzyme kinetic theory, comprised at least three different metastable conformations. In the steady stage, seven conformations, including derivatives from the four conformations in the initial stage, made the reaction pathway complicated. Based on the visualized conformations, we discussed stage-dependent pathways to illustrate the dynamics of the enzyme in action. | |||||||||||||||||||||||||||||||||||||||||||||
History |
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Structure visualization
Supplemental images |
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Downloads & links
-EMDB archive
Map data | ![]() | 26.9 MB | ![]() | |
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Header (meta data) | ![]() ![]() | 21.4 KB 21.4 KB | Display Display | ![]() |
FSC (resolution estimation) | ![]() | 9.2 KB | Display | ![]() |
Images | ![]() | 153.2 KB | ||
Masks | ![]() | 64 MB | ![]() | |
Filedesc metadata | ![]() | 6.6 KB | ||
Others | ![]() ![]() | 60 MB 60 MB | ||
Archive directory | ![]() ![]() | HTTPS FTP |
-Validation report
Summary document | ![]() | 879.4 KB | Display | ![]() |
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Full document | ![]() | 878.9 KB | Display | |
Data in XML | ![]() | 16.1 KB | Display | |
Data in CIF | ![]() | 21.3 KB | Display | |
Arichive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 8xcwMC ![]() 8xcoC ![]() 8xcpC ![]() 8xcqC ![]() 8xcrC ![]() 8xcsC ![]() 8xctC ![]() 8xcuC ![]() 8xcvC ![]() 8xcxC ![]() 8xcyC ![]() 8xczC ![]() 8xd0C ![]() 8xd1C ![]() 8xd2C ![]() 8xd3C ![]() 8xd4C ![]() 8xd5C ![]() 8xd6C M: atomic model generated by this map C: citing same article ( |
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Similar structure data | Similarity search - Function & homology ![]() |
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Links
EMDB pages | ![]() ![]() |
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Map
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Voxel size | X=Y=Z: 0.752 Å | ||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||
Details | EMDB XML:
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-Supplemental data
-Mask #1
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Density Histograms |
-Half map: #2
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Density Histograms |
-Half map: #1
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Density Histograms |
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Sample components
-Entire : Hexamer of glutamate dehydrogenase in the presence of NADP and gl...
Entire | Name: Hexamer of glutamate dehydrogenase in the presence of NADP and glutamate |
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Components |
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-Supramolecule #1: Hexamer of glutamate dehydrogenase in the presence of NADP and gl...
Supramolecule | Name: Hexamer of glutamate dehydrogenase in the presence of NADP and glutamate type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1 |
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Source (natural) | Organism: ![]() ![]() |
Molecular weight | Theoretical: 280 KDa |
-Macromolecule #1: Glutamate dehydrogenase
Macromolecule | Name: Glutamate dehydrogenase / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO / EC number: glutamate dehydrogenase [NAD(P)+] |
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Source (natural) | Organism: ![]() ![]() |
Molecular weight | Theoretical: 46.758477 KDa |
Recombinant expression | Organism: ![]() ![]() |
Sequence | String: MVEIDPFEMA VKQLERAAQY MDISEEALEW LKKPMRIVEV SVPIEMDDGS VKVFTGFRVQ HNWARGPTKG GIRWHPAETL STVKALATW MTWKVAVVDL PYGGGKGGII VNPKELSERE QERLARAYIR AVYDVIGPWT DIPAPDVYTN PKIMGWMMDE Y ETIMRRKG ...String: MVEIDPFEMA VKQLERAAQY MDISEEALEW LKKPMRIVEV SVPIEMDDGS VKVFTGFRVQ HNWARGPTKG GIRWHPAETL STVKALATW MTWKVAVVDL PYGGGKGGII VNPKELSERE QERLARAYIR AVYDVIGPWT DIPAPDVYTN PKIMGWMMDE Y ETIMRRKG PAFGVITGKP LSIGGSLGRG TATAQGAIFT IREAAKALGI DLKGKKIAVQ GYGNAGYYTA KLAKEQLGMT VV AVSDSRG GIYNPDGLDP DEVLKWKREH GSVKDFPGAT NITNEELLEL EVDVLAPAAI EEVITEKNAD NIKAKIVAEV ANG PVTPEA DDILREKGIL QIPDFLCNAG GVTVSYFEWV QNINGYYWTE EEVREKLDKK MTKAFWEVYN THKDKNIHMR DAAY VVAVS RVYQAMKDRG WVKK UniProtKB: Glutamate dehydrogenase |
-Macromolecule #2: NADPH DIHYDRO-NICOTINAMIDE-ADENINE-DINUCLEOTIDE PHOSPHATE
Macromolecule | Name: NADPH DIHYDRO-NICOTINAMIDE-ADENINE-DINUCLEOTIDE PHOSPHATE type: ligand / ID: 2 / Number of copies: 1 / Formula: NDP |
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Molecular weight | Theoretical: 745.421 Da |
Chemical component information | ![]() ChemComp-NDP: |
-Macromolecule #3: 2-OXOGLUTARIC ACID
Macromolecule | Name: 2-OXOGLUTARIC ACID / type: ligand / ID: 3 / Number of copies: 1 / Formula: AKG |
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Molecular weight | Theoretical: 146.098 Da |
Chemical component information | ![]() ChemComp-AKG: |
-Macromolecule #4: water
Macromolecule | Name: water / type: ligand / ID: 4 / Number of copies: 17 / Formula: HOH |
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Molecular weight | Theoretical: 18.015 Da |
Chemical component information | ![]() ChemComp-HOH: |
-Experimental details
-Structure determination
Method | cryo EM |
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![]() | single particle reconstruction |
Aggregation state | particle |
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Sample preparation
Concentration | 4.8 mg/mL | ||||||||||||
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Buffer | pH: 7.5 Component:
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Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 281 K / Instrument: FEI VITROBOT MARK IV Details: The sample solution kept at room temperature was flash-frozen 1-h after mixing GDH, NADP and glutamate solutions.. |
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Electron microscopy
Microscope | JEOL CRYO ARM 300 |
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Details | Grid information as following: Company/model: Quantifoil Cu 1.2/1.3 Material:Cu Grid mesh: 200 |
Image recording | Film or detector model: GATAN K3 (6k x 4k) / Number real images: 7075 / Average electron dose: 1.0 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: ![]() |
Electron optics | Illumination mode: OTHER / Imaging mode: BRIGHT FIELD / Nominal defocus max: 47.0 µm / Nominal defocus min: 0.4 µm |
Sample stage | Cooling holder cryogen: NITROGEN |
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Image processing
-Atomic model buiding 1
Refinement | Protocol: RIGID BODY FIT |
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Output model | ![]() PDB-8xcw: |