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- PDB-9m2q: Imidazole glycerol phosphate dehydratase from Mycobacterium tuber... -

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Basic information

Entry
Database: PDB / ID: 9m2q
TitleImidazole glycerol phosphate dehydratase from Mycobacterium tuberculosis, in complex with aminotriazole
ComponentsImidazoleglycerol-phosphate dehydratase
KeywordsLYASE / histidine pathway / dehydratase / Mycobacterium tuberculosis
Function / homology
Function and homology information


imidazoleglycerol-phosphate dehydratase / imidazoleglycerol-phosphate dehydratase activity / L-histidine biosynthetic process / metal ion binding / cytoplasm
Similarity search - Function
Imidazoleglycerol-phosphate dehydratase signature 1. / Imidazoleglycerol-phosphate dehydratase / Imidazoleglycerol-phosphate dehydratase, conserved site / Imidazole glycerol phosphate dehydratase domain superfamily / Imidazoleglycerol-phosphate dehydratase / Imidazoleglycerol-phosphate dehydratase signature 2. / Ribosomal protein S5 domain 2-type fold
Similarity search - Domain/homology
3-AMINO-1,2,4-TRIAZOLE / : / Imidazoleglycerol-phosphate dehydratase
Similarity search - Component
Biological speciesMycobacterium tuberculosis (bacteria)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.8 Å
AuthorsRaina, R. / Kar, D. / Singla, M. / Tiwari, S. / Kumari, S. / Aneja, S. / Kumar, V. / Banerjee, S. / Goyal, S. / Pal, R.K. ...Raina, R. / Kar, D. / Singla, M. / Tiwari, S. / Kumari, S. / Aneja, S. / Kumar, V. / Banerjee, S. / Goyal, S. / Pal, R.K. / Vinothkumar, K.R. / Biswal, B.K.
Funding support India, 4items
OrganizationGrant numberCountry
Department of Biotechnology (DBT, India)BT/HRD/TIF/09/02/2021-22 India
Department of Biotechnology (DBT, India)BT/PR29075/BRB/10/1699/2018 India
Department of Biotechnology (DBT, India)BT/PR40325/BTIS/137/1/2020 India
Department of Biotechnology (DBT, India)DBT/PR12422/MED/31/287/2014 India
CitationJournal: Acta Crystallogr F Struct Biol Commun / Year: 2025
Title: Cryo-EM structures of Mycobacterium tuberculosis imidazole glycerol phosphate dehydratase in the apo state and in the presence of small molecules.
Authors: Rahul Raina / Deepsikha Kar / Mohini Singla / Satish Tiwari / Swati Kumari / Sonanjali Aneja / Varun Kumar / Soumya Banerjee / Shivika Goyal / Ravi Kant Pal / Kutti R Vinothkumar / Bichitra Biswal /
Abstract: Unlike humans, Mycobacterium tuberculosis (Mtb), the causative agent of human tuberculosis, has a de novo histidine-biosynthesis pathway. The enzyme imidazole glycerol phosphate dehydratase (IGPD), ...Unlike humans, Mycobacterium tuberculosis (Mtb), the causative agent of human tuberculosis, has a de novo histidine-biosynthesis pathway. The enzyme imidazole glycerol phosphate dehydratase (IGPD), which catalyses the conversion of imidazole glycerol phosphate to imidazole acetol phosphate, has been studied extensively from various organisms and has become a major target for the development of antibacterial, antiweed and antifungal small molecules. In our previous studies, we have shown that in crystals IGPD forms a 24-mer oligomeric state in which the monomers are arranged in 432 symmetry. In order to gain insights into the oligomeric state of Mtb IGPD in solution, we determined cryogenic sample electron microscopy (cryo-EM) structures of apo IGPD at 2.2 and 3.1 Å resolution. In addition, we also determined the cryo-EM structure of IGPD in the presence of 3-amino-1,2,4-triazole (ATZ) to 2.8 Å resolution. The results of this work, which corroborate those from the crystallographic studies, indicate that IGPD forms a homo-oligomeric structure in solution comprising of 24 subunits. ATZ binds in the active-site pocket of the enzyme, which is located at the interface of three monomers and tethers 24 ATZ molecules. The results of this study suggest that cryo-EM, in addition to being a rapidly evolving and complementary imaging technology for elucidating 3D structures of biological macromolecules, can be useful in pinpointing the mode of binding small molecules of low mass (here ∼85 Da) and mapping protein-ligand interactions, which could assist in the design of accurate (high-potency) inhibitors.
History
DepositionFeb 28, 2025Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0Jun 18, 2025Provider: repository / Type: Initial release

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Imidazoleglycerol-phosphate dehydratase
B: Imidazoleglycerol-phosphate dehydratase
C: Imidazoleglycerol-phosphate dehydratase
D: Imidazoleglycerol-phosphate dehydratase
E: Imidazoleglycerol-phosphate dehydratase
F: Imidazoleglycerol-phosphate dehydratase
G: Imidazoleglycerol-phosphate dehydratase
H: Imidazoleglycerol-phosphate dehydratase
I: Imidazoleglycerol-phosphate dehydratase
J: Imidazoleglycerol-phosphate dehydratase
K: Imidazoleglycerol-phosphate dehydratase
L: Imidazoleglycerol-phosphate dehydratase
M: Imidazoleglycerol-phosphate dehydratase
N: Imidazoleglycerol-phosphate dehydratase
O: Imidazoleglycerol-phosphate dehydratase
P: Imidazoleglycerol-phosphate dehydratase
Q: Imidazoleglycerol-phosphate dehydratase
R: Imidazoleglycerol-phosphate dehydratase
S: Imidazoleglycerol-phosphate dehydratase
T: Imidazoleglycerol-phosphate dehydratase
U: Imidazoleglycerol-phosphate dehydratase
V: Imidazoleglycerol-phosphate dehydratase
W: Imidazoleglycerol-phosphate dehydratase
X: Imidazoleglycerol-phosphate dehydratase
hetero molecules


Theoretical massNumber of molelcules
Total (without water)571,85996
Polymers567,20424
Non-polymers4,65572
Water00
1


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

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Components

#1: Protein ...
Imidazoleglycerol-phosphate dehydratase / IGPD


Mass: 23633.516 Da / Num. of mol.: 24
Source method: isolated from a genetically manipulated source
Details: The enzyme was expressed with a Histag at the N-terminus but not observed in the experiment. The C-terminal residues are also not observed in the experiment. Uniparc ID is UPI000012C861.
Source: (gene. exp.) Mycobacterium tuberculosis (bacteria) / Gene: hisB, Rv1601, MTCY336.03c / Production host: Mycolicibacterium smegmatis (bacteria) / Strain (production host): MC2 (4517)
References: UniProt: P9WML9, imidazoleglycerol-phosphate dehydratase
#2: Chemical...
ChemComp-MN / MANGANESE (II) ION


Mass: 54.938 Da / Num. of mol.: 48 / Source method: obtained synthetically / Formula: Mn / Feature type: SUBJECT OF INVESTIGATION
#3: Chemical...
ChemComp-3TR / 3-AMINO-1,2,4-TRIAZOLE / AMITROLE


Mass: 84.080 Da / Num. of mol.: 24 / Source method: obtained synthetically / Formula: C2H4N4 / Feature type: SUBJECT OF INVESTIGATION / Comment: inhibitor*YM
Has ligand of interestY
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: Mycobacterium tuberculosis HisB with 3TR / Type: COMPLEX / Details: An oligomer of 24 subunits / Entity ID: #1 / Source: RECOMBINANT
Molecular weightValue: 0.55 MDa / Experimental value: NO
Source (natural)Organism: Mycobacterium tuberculosis (bacteria)
Source (recombinant)Organism: Mycolicibacterium smegmatis (bacteria) / Strain: MC2 (4517)
Buffer solutionpH: 8.5
Buffer component
IDConc.NameFormulaBuffer-ID
120 mMTris1
2200 mMSodium ChlorideNaCl1
SpecimenConc.: 1.2 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Details: Enzyme was incubated with 10 fold excess of 3TR and this mixture was directly used for freezing
Specimen supportGrid material: GOLD / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R0.6/1
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 291 K

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: TFS KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal magnification: 75000 X / Calibrated magnification: 130841 X / Nominal defocus max: 35000 nm / Nominal defocus min: 12000 nm / Cs: 2.7 mm / C2 aperture diameter: 50 µm / Alignment procedure: COMA FREE
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingAverage exposure time: 60 sec. / Electron dose: 27.7 e/Å2 / Detector mode: COUNTING / Film or detector model: FEI FALCON III (4k x 4k) / Num. of grids imaged: 1
Image scansSampling size: 14 µm / Width: 4096 / Height: 4096

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Processing

EM software
IDNameVersionCategory
1RELIONparticle selection
2EPUimage acquisition
4CTFFIND4.1CTF correction
5RELIONCTF correction
8UCSF Chimera1.18model fitting
9Coot0.9.8.95model fitting
11RELIONinitial Euler assignment
12RELIONfinal Euler assignment
13RELIONclassification
14RELION3D reconstruction
15REFMAC5.8.0430model refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 189390
SymmetryPoint symmetry: O (octahedral)
3D reconstructionResolution: 2.8 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 23789 / Algorithm: BACK PROJECTION / Num. of class averages: 1 / Symmetry type: POINT
Atomic model buildingB value: 133.9 / Protocol: OTHER / Space: RECIPROCAL
Atomic model buildingPDB-ID: 4LPF

4lpf


Pdb chain-ID: A / Accession code: 4LPF / Chain residue range: 10-199 / Pdb chain residue range: 10-199 / Source name: PDB / Type: experimental model
RefinementResolution: 2.8→141.24 Å / Cor.coef. Fo:Fc: 0.949 / SU B: 8.022 / SU ML: 0.155 / ESU R: 0.422
Stereochemistry target values: MAXIMUM LIKELIHOOD WITH PHASES
Details: HYDROGENS HAVE BEEN USED IF PRESENT IN THE INPUT
RfactorNum. reflection% reflection
Rwork0.28474 --
obs0.28474 268676 100 %
Solvent computationSolvent model: PARAMETERS FOR MASK CACLULATION
Displacement parametersBiso mean: 131.451 Å2
Refinement stepCycle: 1 / Total: 35448
Refine LS restraints
Refine-IDTypeDev idealDev ideal targetNumber
ELECTRON MICROSCOPYr_bond_refined_d0.0030.01236120
ELECTRON MICROSCOPYr_bond_other_d00.01633624
ELECTRON MICROSCOPYr_angle_refined_deg0.9821.82149080
ELECTRON MICROSCOPYr_angle_other_deg0.3621.77376872
ELECTRON MICROSCOPYr_dihedral_angle_1_deg5.62554536
ELECTRON MICROSCOPYr_dihedral_angle_2_deg3.6135432
ELECTRON MICROSCOPYr_dihedral_angle_3_deg11.609105592
ELECTRON MICROSCOPYr_dihedral_angle_4_deg
ELECTRON MICROSCOPYr_chiral_restr0.0460.25592
ELECTRON MICROSCOPYr_gen_planes_refined0.0030.0244064
ELECTRON MICROSCOPYr_gen_planes_other0.0010.028640
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_it5.59212.59418216
ELECTRON MICROSCOPYr_mcbond_other5.59212.59418216
ELECTRON MICROSCOPYr_mcangle_it8.62922.65922728
ELECTRON MICROSCOPYr_mcangle_other8.62922.6622729
ELECTRON MICROSCOPYr_scbond_it7.72513.84917904
ELECTRON MICROSCOPYr_scbond_other7.72513.8517905
ELECTRON MICROSCOPYr_scangle_it
ELECTRON MICROSCOPYr_scangle_other12.92324.73726353
ELECTRON MICROSCOPYr_long_range_B_refined17.952151.99145222
ELECTRON MICROSCOPYr_long_range_B_other17.952151.99145223
ELECTRON MICROSCOPYr_rigid_bond_restr
ELECTRON MICROSCOPYr_sphericity_free
ELECTRON MICROSCOPYr_sphericity_bonded
LS refinement shellResolution: 2.801→2.874 Å / Total num. of bins used: 20
RfactorNum. reflection% reflection
Rfree0 0 -
Rwork1.324 19945 -
obs--100 %

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