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- PDB-8z1n: Cryo-EM structure of apo Aspergillus terreus glutamate dehydrogen... -

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Basic information

Entry
Database: PDB / ID: 8z1n
TitleCryo-EM structure of apo Aspergillus terreus glutamate dehydrogenase (AtGDH) in the closed conformation (form 2)
ComponentsGlutamate dehydrogenase
KeywordsOXIDOREDUCTASE / glutamate dehydrogenase / allostery / cooperativity / Aspergillus / cryo-EM / domain dynamics
Function / homology
Function and homology information


glutamate biosynthetic process / glutamate dehydrogenase (NADP+) activity / nucleotide binding / cytosol
Similarity search - Function
: / Glutamate dehydrogenase / NAD(P) binding domain of glutamate dehydrogenase / Leu/Phe/Val dehydrogenases active site / Glu / Leu / Phe / Val dehydrogenases active site. / Glutamate/phenylalanine/leucine/valine dehydrogenase / Glutamate/phenylalanine/leucine/valine dehydrogenase, dimerisation domain / Glu/Leu/Phe/Val dehydrogenase, dimerisation domain / Glutamate/Leucine/Phenylalanine/Valine dehydrogenase / Glutamate/phenylalanine/leucine/valine dehydrogenase, C-terminal ...: / Glutamate dehydrogenase / NAD(P) binding domain of glutamate dehydrogenase / Leu/Phe/Val dehydrogenases active site / Glu / Leu / Phe / Val dehydrogenases active site. / Glutamate/phenylalanine/leucine/valine dehydrogenase / Glutamate/phenylalanine/leucine/valine dehydrogenase, dimerisation domain / Glu/Leu/Phe/Val dehydrogenase, dimerisation domain / Glutamate/Leucine/Phenylalanine/Valine dehydrogenase / Glutamate/phenylalanine/leucine/valine dehydrogenase, C-terminal / Glutamate/Leucine/Phenylalanine/Valine dehydrogenase / Aminoacid dehydrogenase-like, N-terminal domain superfamily / NAD(P)-binding domain superfamily
Similarity search - Domain/homology
Glutamate dehydrogenase
Similarity search - Component
Biological speciesAspergillus terreus (mold)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.18 Å
AuthorsGodsora, B.K.J. / Das, P. / Bhaumik, P.
Funding support India, 1items
OrganizationGrant numberCountry
Science and Engineering Research Board (SERB)CRG/2021/002404 India
CitationJournal: Protein Sci / Year: 2025
Title: Conformational flexibility associated with remote residues regulates the kinetic properties of glutamate dehydrogenase.
Authors: Barsa Kanchan Jyotshna Godsora / Parijat Das / Prasoon Kumar Mishra / Anjali Sairaman / Sandip Kaledhonkar / Narayan S Punekar / Prasenjit Bhaumik /
Abstract: Glutamate dehydrogenase (GDH) is a pivotal metabolic enzyme in all living organisms, and some of the GDHs exhibit substrate-dependent homotropic cooperativity. However, the mode of allosteric ...Glutamate dehydrogenase (GDH) is a pivotal metabolic enzyme in all living organisms, and some of the GDHs exhibit substrate-dependent homotropic cooperativity. However, the mode of allosteric communication during the homotropic effect in GDHs remains poorly understood. In this study, we examined two homologous GDHs, Aspergillus niger GDH (AnGDH) and Aspergillus terreus GDH (AtGDH), with differing substrate utilization kinetics to uncover the factors driving their distinct behavior. We report the crystal structures and first-ever cryo-EM structures of apo- AtGDH and AnGDH that captured arrays of conformational ensembles. A wider mouth opening (~ 21 Å) is observed for the cooperative AnGDH as compared to the non-cooperative AtGDH (~17 Å) in their apo states. A network of interactions related to the substitutions in Domain II influence structural flexibility in these GDHs. Remarkably, we have identified a distant substitution (R246 to S) in Domain II, as a part of this network, which can impact the mouth opening and converts non-cooperative AtGDH into a cooperative enzyme. Our study demonstrates that remote residues can influence structural and kinetic properties in homologous GDHs.
History
DepositionApr 11, 2024Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0Mar 5, 2025Provider: repository / Type: Initial release
Revision 1.0Mar 5, 2025Data content type: EM metadata / Data content type: EM metadata / Provider: repository / Type: Initial release
Revision 1.0Mar 5, 2025Data content type: Half map / Part number: 1 / Data content type: Half map / Provider: repository / Type: Initial release
Revision 1.0Mar 5, 2025Data content type: Half map / Part number: 2 / Data content type: Half map / Provider: repository / Type: Initial release
Revision 1.0Mar 5, 2025Data content type: Image / Data content type: Image / Provider: repository / Type: Initial release
Revision 1.0Mar 5, 2025Data content type: Primary map / Data content type: Primary map / Provider: repository / Type: Initial release
Revision 1.1Jul 2, 2025Group: Data collection / Category: em_admin / em_software / Item: _em_admin.last_update
Revision 1.1Jul 2, 2025Data content type: EM metadata / Data content type: EM metadata / EM metadata / Group: Data processing / Experimental summary / Data content type: EM metadata / EM metadata / Category: em_admin / em_software / Data content type: EM metadata / Item: _em_admin.last_update

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
D: Glutamate dehydrogenase
E: Glutamate dehydrogenase
F: Glutamate dehydrogenase
B: Glutamate dehydrogenase
C: Glutamate dehydrogenase
A: Glutamate dehydrogenase


Theoretical massNumber of molelcules
Total (without water)295,4856
Polymers295,4856
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: gel filtration, not applicable
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: Protein
Glutamate dehydrogenase


Mass: 49247.438 Da / Num. of mol.: 6
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Aspergillus terreus (mold) / Gene: gdhA, ATETN484_0007063400 / Production host: Escherichia coli (E. coli) / References: UniProt: T2D1F5
Has protein modificationN

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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: apo form of Aspergillus terreus glutamate dehydrogenase
Type: COMPLEX / Entity ID: all / Source: RECOMBINANT
Molecular weightValue: 0.3 MDa / Experimental value: YES
Source (natural)Organism: Aspergillus terreus (mold)
Source (recombinant)Organism: Escherichia coli (E. coli)
Buffer solutionpH: 7.5 / Details: 30 mM Phosphate buffer pH 7.5
SpecimenConc.: 3 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: GOLD / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R0.6/1
VitrificationCryogen name: ETHANE

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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: SPOT SCAN
Electron lensMode: BRIGHT FIELD / Nominal defocus max: 3300 nm / Nominal defocus min: 2100 nm
Image recordingElectron dose: 25.75 e/Å2 / Detector mode: COUNTING / Film or detector model: FEI FALCON III (4k x 4k)

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Processing

EM software
IDNameVersionCategory
1REFMAC5.8.0352model refinement
2PHENIXmodel refinement
CTF correctionType: NONE
SymmetryPoint symmetry: D3 (2x3 fold dihedral)
3D reconstructionResolution: 3.18 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 81516 / Num. of class averages: 1 / Symmetry type: POINT
RefinementResolution: 3.18→3.18 Å / Cor.coef. Fo:Fc: 0.961 / SU B: 19.058 / SU ML: 0.329 / ESU R: 0.721
Stereochemistry target values: MAXIMUM LIKELIHOOD WITH PHASES
Details: HYDROGENS HAVE BEEN ADDED IN THE RIDING POSITIONS
RfactorNum. reflection% reflection
Rwork0.33621 --
obs0.33621 128025 100 %
Solvent computationSolvent model: PARAMETERS FOR MASK CACLULATION
Displacement parametersBiso mean: 175.767 Å2
Refinement stepCycle: 1 / Total: 20789
Refine LS restraints
Refine-IDTypeDev idealDev ideal targetNumber
ELECTRON MICROSCOPYr_bond_refined_d0.0070.01221208
ELECTRON MICROSCOPYr_bond_other_d00.01619350
ELECTRON MICROSCOPYr_angle_refined_deg1.1791.63228697
ELECTRON MICROSCOPYr_angle_other_deg0.4291.5645012
ELECTRON MICROSCOPYr_dihedral_angle_1_deg4.55952754
ELECTRON MICROSCOPYr_dihedral_angle_2_deg4.8675120
ELECTRON MICROSCOPYr_dihedral_angle_3_deg18.185103434
ELECTRON MICROSCOPYr_dihedral_angle_4_deg
ELECTRON MICROSCOPYr_chiral_restr0.0550.23119
ELECTRON MICROSCOPYr_gen_planes_refined0.0050.0224732
ELECTRON MICROSCOPYr_gen_planes_other0.0010.024292
ELECTRON MICROSCOPYr_nbd_refined
ELECTRON MICROSCOPYr_nbd_other
ELECTRON MICROSCOPYr_nbtor_refined
ELECTRON MICROSCOPYr_nbtor_other
ELECTRON MICROSCOPYr_xyhbond_nbd_refined
ELECTRON MICROSCOPYr_xyhbond_nbd_other
ELECTRON MICROSCOPYr_metal_ion_refined
ELECTRON MICROSCOPYr_metal_ion_other
ELECTRON MICROSCOPYr_symmetry_vdw_refined
ELECTRON MICROSCOPYr_symmetry_vdw_other
ELECTRON MICROSCOPYr_symmetry_hbond_refined
ELECTRON MICROSCOPYr_symmetry_hbond_other
ELECTRON MICROSCOPYr_symmetry_metal_ion_refined
ELECTRON MICROSCOPYr_symmetry_metal_ion_other
ELECTRON MICROSCOPYr_mcbond_it16.06117.59111034
ELECTRON MICROSCOPYr_mcbond_other16.0617.59111034
ELECTRON MICROSCOPYr_mcangle_it23.5326.37113782
ELECTRON MICROSCOPYr_mcangle_other23.5326.37113783
ELECTRON MICROSCOPYr_scbond_it15.62518.41210174
ELECTRON MICROSCOPYr_scbond_other15.62418.41210175
ELECTRON MICROSCOPYr_scangle_it
ELECTRON MICROSCOPYr_scangle_other23.44727.1914916
ELECTRON MICROSCOPYr_long_range_B_refined35.20690477
ELECTRON MICROSCOPYr_long_range_B_other35.20690478
ELECTRON MICROSCOPYr_rigid_bond_restr
ELECTRON MICROSCOPYr_sphericity_free
ELECTRON MICROSCOPYr_sphericity_bonded
LS refinement shellResolution: 3.15→3.232 Å / Total num. of bins used: 20
RfactorNum. reflection% reflection
Rfree0 0 -
Rwork1.014 9477 -
obs--100 %

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