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- PDB-9i0k: Mycobacterium smegmatis inosine monophosphate dehydrogenase (IMPD... -

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Basic information

Entry
Database: PDB / ID: 9i0k
TitleMycobacterium smegmatis inosine monophosphate dehydrogenase (IMPDH) E-XMP* intermediate, compressed
ComponentsInosine-5'-monophosphate dehydrogenase
KeywordsOXIDOREDUCTASE / Octamer / reaction intermediate / Purine metabolism / IMPDH
Function / homology
Function and homology information


IMP dehydrogenase / IMP dehydrogenase activity / GMP biosynthetic process / GTP biosynthetic process / nucleotide binding / metal ion binding
Similarity search - Function
IMP dehydrogenase / GMP reductase, conserved site / IMP dehydrogenase / GMP reductase signature. / Inosine-5'-monophosphate dehydrogenase / IMP dehydrogenase/GMP reductase / IMP dehydrogenase / GMP reductase domain / IMP dehydrogenase / GMP reductase domain / Domain in cystathionine beta-synthase and other proteins. / CBS domain superfamily / CBS domain / CBS domain ...IMP dehydrogenase / GMP reductase, conserved site / IMP dehydrogenase / GMP reductase signature. / Inosine-5'-monophosphate dehydrogenase / IMP dehydrogenase/GMP reductase / IMP dehydrogenase / GMP reductase domain / IMP dehydrogenase / GMP reductase domain / Domain in cystathionine beta-synthase and other proteins. / CBS domain superfamily / CBS domain / CBS domain / CBS domain profile. / Aldolase-type TIM barrel
Similarity search - Domain/homology
INOSINIC ACID / : / NICOTINAMIDE-ADENINE-DINUCLEOTIDE / Inosine-5'-monophosphate dehydrogenase
Similarity search - Component
Biological speciesMycolicibacterium smegmatis MC2 155 (bacteria)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.73 Å
AuthorsBulvas, O. / Kouba, T. / Pichova, I.
Funding supportEuropean Union, 1items
OrganizationGrant numberCountry
European Union (EU)LX22NPO5103European Union
CitationJournal: J Struct Biol / Year: 2025
Title: Conformational landscape of the mycobacterial inosine 5'-monophosphate dehydrogenase octamerization interface.
Authors: Ondřej Bulvas / Zdeněk Knejzlík / Anatolij Filimoněnko / Tomáš Kouba / Iva Pichová /
Abstract: Inosine 5'-monophosphate dehydrogenase (IMPDH), a key enzyme in bacterial purine metabolism, plays an essential role in the biosynthesis of guanine nucleotides and shows promise as a target for ...Inosine 5'-monophosphate dehydrogenase (IMPDH), a key enzyme in bacterial purine metabolism, plays an essential role in the biosynthesis of guanine nucleotides and shows promise as a target for antimicrobial drug development. Despite its significance, the conformational dynamics and substrate-induced structural changes in bacterial IMPDH remain poorly understood, particularly with respect to its octameric assembly. Using cryo-EM, we present full-length structures of IMPDH from Mycobacterium smegmatis (MsmIMPDH) captured in a reaction intermediate state, revealing conformational changes upon substrate binding. The structures feature resolved flexible loops that coordinate the binding of the substrate, the cofactor, and the K ion. Our structural analysis identifies a novel octamerization interface unique to MsmIMPDH. Additionally, a previously unobserved barrel-like density suggests potential self-interactions within the C-terminal regions, hinting at a regulatory mechanism tied to assembly and function of the enzyme. These data provide insights into substrate-induced conformational dynamics and novel interaction interfaces in MsmIMPDH, potentially informing the development of IMPDH-targeted drugs.
History
DepositionJan 15, 2025Deposition site: PDBE / Processing site: PDBE
Revision 1.0Mar 26, 2025Provider: repository / Type: Initial release
Revision 1.1Apr 2, 2025Group: Data collection / Database references / Category: citation / em_admin
Item: _citation.journal_volume / _citation.page_last ..._citation.journal_volume / _citation.page_last / _citation.pdbx_database_id_PubMed / _citation.title / _em_admin.last_update

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Inosine-5'-monophosphate dehydrogenase
B: Inosine-5'-monophosphate dehydrogenase
C: Inosine-5'-monophosphate dehydrogenase
D: Inosine-5'-monophosphate dehydrogenase
E: Inosine-5'-monophosphate dehydrogenase
F: Inosine-5'-monophosphate dehydrogenase
G: Inosine-5'-monophosphate dehydrogenase
H: Inosine-5'-monophosphate dehydrogenase
hetero molecules


Theoretical massNumber of molelcules
Total (without water)435,51832
Polymers427,1128
Non-polymers8,40624
Water1,44180
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: electron microscopy
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1

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Components

#1: Protein
Inosine-5'-monophosphate dehydrogenase / IMP dehydrogenase / IMPD / IMPDH


Mass: 53388.988 Da / Num. of mol.: 8
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Mycolicibacterium smegmatis MC2 155 (bacteria)
Strain: MC2 155 / Gene: guaB, MSMEG_1602 / Plasmid: pRSFDuet-1 / Production host: Escherichia coli BL21(DE3) (bacteria) / Strain (production host): BL21(DE3) / Variant (production host): LOBSTR- -RIL / References: UniProt: A0QSU3, IMP dehydrogenase
#2: Chemical
ChemComp-IMP / INOSINIC ACID


Mass: 348.206 Da / Num. of mol.: 8 / Source method: obtained synthetically / Formula: C10H13N4O8P
#3: Chemical
ChemComp-NAD / NICOTINAMIDE-ADENINE-DINUCLEOTIDE


Mass: 663.425 Da / Num. of mol.: 8 / Source method: obtained synthetically / Formula: C21H27N7O14P2 / Comment: NAD*YM
#4: Chemical
ChemComp-K / POTASSIUM ION


Mass: 39.098 Da / Num. of mol.: 8 / Source method: obtained synthetically / Formula: K
#5: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 80 / Source method: isolated from a natural source / Formula: H2O
Has ligand of interestN
Has protein modificationY

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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: Octameric assembly of inosine monophosphate dehydrogenase reaction intermediate; E-XMP* in complex with NAD+
Type: COMPLEX / Entity ID: #1 / Source: RECOMBINANT
Molecular weightExperimental value: NO
Source (natural)Organism: Mycolicibacterium smegmatis MC2 155 (bacteria)
Source (recombinant)Organism: Escherichia coli BL21(DE3) (bacteria) / Strain: LOBSTR-BL21(DE3)-RIL / Plasmid: pRSFDuet-1
Buffer solutionpH: 7.5
Details: 50 mM HEPES (pH 7.5), 200 mM KCl, 5 mM DTT, 32 mM MgCl2 Ligand: 10 mM ATP + 10 mM IMP + 10 mM NAD
Buffer component
IDConc.NameFormulaBuffer-ID
150 mM2-[4-(2-Hydroxyethyl)piperazin-1-yl]ethane-1-sulfonic acidHEPES1
2200 mMPotassium chlorideKCl1
35 mMDithiothreitolDTT1
432 mMMagnesium chlorideMgCl21
510 mMInosine monophosphateIMP1
610 mMNicotinamide adenine dinucleotideNAD1
710 mMAdenosine triphosphateATP1
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: GOLD / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R2/1
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 %

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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: 165000 X / Nominal defocus max: 3200 nm / Nominal defocus min: 1200 nm / Cs: 2.7 mm / Alignment procedure: ZEMLIN TABLEAU
Specimen holderCryogen: NITROGEN
Image recordingAverage exposure time: 2 sec. / Electron dose: 40.1 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Num. of real images: 8691

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Processing

EM softwareName: PHENIX / Version: 1.21rc1_5084 / Category: model refinement
CTF correctionType: PHASE FLIPPING ONLY
Particle selectionNum. of particles selected: 4833002
3D reconstructionResolution: 2.73 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 124968 / Symmetry type: POINT
Atomic model buildingB value: 37.48 / Protocol: RIGID BODY FIT / Target criteria: CC coefficient
Details: Initial fitting was done in UCSF ChimeraX. Model refinement was done by iterative cycles of manual fitting with Coot and ISOLDE and automated fitting with phenix.real_space_refine.
Atomic model buildingPDB-ID: 8QQV

8qqv


Accession code: 8QQV / Source name: PDB / Type: experimental model
RefinementCross valid method: NONE
Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2
Displacement parametersBiso mean: 38.75 Å2
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.007721656
ELECTRON MICROSCOPYf_angle_d0.640729536
ELECTRON MICROSCOPYf_chiral_restr0.05473584
ELECTRON MICROSCOPYf_plane_restr0.00443840
ELECTRON MICROSCOPYf_dihedral_angle_d11.94867864

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