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Yorodumi- PDB-7a1d: Cryo-EM map of the large glutamate dehydrogenase composed of 180 ... -
+Open data
-Basic information
Entry | Database: PDB / ID: 7a1d | ||||||
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Title | Cryo-EM map of the large glutamate dehydrogenase composed of 180 kDa subunits from Mycobacterium smegmatis (open conformation) | ||||||
Components | NAD-specific glutamate dehydrogenase | ||||||
Keywords | OXIDOREDUCTASE / large glutamate dehydrogenase Mycobacterium metabolism | ||||||
Function / homology | Function and homology information | ||||||
Biological species | Mycolicibacterium smegmatis (bacteria) | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 4.19 Å | ||||||
Authors | Lazaro, M. / Melero, R. / Huet, C. / Lopez-Alonso, J.P. / Delgado, S. / Dodu, A. / Bruch, E.M. / Abriata, L.A. / Alzari, P.M. / Valle, M. / Lisa, M.N. | ||||||
Funding support | Spain, 1items
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Citation | Journal: Commun Biol / Year: 2021 Title: 3D architecture and structural flexibility revealed in the subfamily of large glutamate dehydrogenases by a mycobacterial enzyme. Authors: Melisa Lázaro / Roberto Melero / Charlotte Huet / Jorge P López-Alonso / Sandra Delgado / Alexandra Dodu / Eduardo M Bruch / Luciano A Abriata / Pedro M Alzari / Mikel Valle / María-Natalia Lisa / Abstract: Glutamate dehydrogenases (GDHs) are widespread metabolic enzymes that play key roles in nitrogen homeostasis. Large glutamate dehydrogenases composed of 180 kDa subunits (L-GDHs) contain long N- ...Glutamate dehydrogenases (GDHs) are widespread metabolic enzymes that play key roles in nitrogen homeostasis. Large glutamate dehydrogenases composed of 180 kDa subunits (L-GDHs) contain long N- and C-terminal segments flanking the catalytic core. Despite the relevance of L-GDHs in bacterial physiology, the lack of structural data for these enzymes has limited the progress of functional studies. Here we show that the mycobacterial L-GDH (mL-GDH) adopts a quaternary structure that is radically different from that of related low molecular weight enzymes. Intersubunit contacts in mL-GDH involve a C-terminal domain that we propose as a new fold and a flexible N-terminal segment comprising ACT-like and PAS-type domains that could act as metabolic sensors for allosteric regulation. These findings uncover unique aspects of the structure-function relationship in the subfamily of L-GDHs. | ||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | Molecule: MolmilJmol/JSmol |
-Downloads & links
-Download
PDBx/mmCIF format | 7a1d.cif.gz | 776.7 KB | Display | PDBx/mmCIF format |
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PDB format | pdb7a1d.ent.gz | 596.4 KB | Display | PDB format |
PDBx/mmJSON format | 7a1d.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 7a1d_validation.pdf.gz | 1.2 MB | Display | wwPDB validaton report |
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Full document | 7a1d_full_validation.pdf.gz | 1.2 MB | Display | |
Data in XML | 7a1d_validation.xml.gz | 120.3 KB | Display | |
Data in CIF | 7a1d_validation.cif.gz | 182 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/a1/7a1d ftp://data.pdbj.org/pub/pdb/validation_reports/a1/7a1d | HTTPS FTP |
-Related structure data
Related structure data | 11606MC 7jsrC M: map data used to model this data C: citing same article (ref.) |
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Similar structure data |
-Links
-Assembly
Deposited unit |
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-Components
#1: Protein | Mass: 176341.859 Da / Num. of mol.: 4 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Mycolicibacterium smegmatis (strain ATCC 700084 / mc(2)155) (bacteria) Strain: ATCC 700084 / mc(2)155 / Gene: gdh, MSMEG_4699, MSMEI_4582 / Production host: Escherichia coli BL21 (bacteria) / References: UniProt: A0R1C2, glutamate dehydrogenase |
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-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
-Sample preparation
Component | Name: Cryo-EM map of the large glutamate dehydrogenase composed of 180 kDa subunits from Mycobacterium smegmatis (open conformation) Type: COMPLEX / Entity ID: all / Source: RECOMBINANT |
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Molecular weight | Value: 0.705 MDa / Experimental value: NO |
Source (natural) | Organism: Mycolicibacterium smegmatis MC2 155 (bacteria) |
Source (recombinant) | Organism: Escherichia coli BL21 (bacteria) / Strain: BL21(DE3) |
Buffer solution | pH: 6 |
Specimen | Conc.: 0.3 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES |
Specimen support | Grid material: COPPER / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil |
Vitrification | Cryogen name: ETHANE |
-Electron microscopy imaging
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
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Microscopy | Model: FEI TITAN KRIOS |
Electron gun | Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM |
Electron lens | Mode: BRIGHT FIELD / Calibrated magnification: 47170 X / Calibrated defocus min: 670 nm / Calibrated defocus max: 3260 nm / Cs: 2.7 mm |
Image recording | Electron dose: 40 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k) |
Image scans | Movie frames/image: 20 / Used frames/image: 1-20 |
-Processing
Software |
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EM software |
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CTF correction | Type: PHASE FLIPPING ONLY | ||||||||||||||||||||||||
Symmetry | Point symmetry: D2 (2x2 fold dihedral) | ||||||||||||||||||||||||
3D reconstruction | Resolution: 4.19 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 63715 / Algorithm: FOURIER SPACE / Symmetry type: POINT | ||||||||||||||||||||||||
Atomic model building | Protocol: OTHER Details: Model fitting into the cryo-EM map was performed using the programs UCSF Chimera (Pettersen et al., 2004), Namdinator (Kidmose et al., 2019), phenix.real_space_refine (Afonine et al., 2018) ...Details: Model fitting into the cryo-EM map was performed using the programs UCSF Chimera (Pettersen et al., 2004), Namdinator (Kidmose et al., 2019), phenix.real_space_refine (Afonine et al., 2018) and Coot (Emsley, Lohkamp, Scott, & Cowtan, 2010). Residues 500-1588 from the crystal structure of Se-Met mL-GDH180 (PDB code 7JSR) were fitted into the cryo-EM map. Se-methionine residues were replaced by methionine residues using Coot (Emsley et al., 2010) and the model was finally refined employing phenix.real_space_refine (Afonine et al., 2018) with NCS and secondary structure restraints. | ||||||||||||||||||||||||
Refinement | Stereochemistry target values: CDL v1.2 | ||||||||||||||||||||||||
Refine LS restraints |
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