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- PDB-9gml: Cryo-EM structure of Sporosarcina pasteurii urease -

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Basic information

Entry
Database: PDB / ID: 9gml
TitleCryo-EM structure of Sporosarcina pasteurii urease
Components(Urease subunit ...) x 3
KeywordsHYDROLASE / Urease / enzyme / Nickel / urea
Function / homology
Function and homology information


urease complex / urease / urease activity / urea catabolic process / nickel cation binding / cytoplasm
Similarity search - Function
Urease, gamma subunit / : / Urease active site / Urease active site. / Urease nickel binding site / Urease nickel ligands signature. / Urease, beta subunit / Urease, alpha subunit / Urease alpha subunit, C-terminal / Urease, beta subunit superfamily ...Urease, gamma subunit / : / Urease active site / Urease active site. / Urease nickel binding site / Urease nickel ligands signature. / Urease, beta subunit / Urease, alpha subunit / Urease alpha subunit, C-terminal / Urease, beta subunit superfamily / : / Urease beta subunit / Urease domain profile. / Urease alpha-subunit, N-terminal domain / Urease alpha-subunit, N-terminal domain / Urease, gamma/gamma-beta subunit / Urease, gamma subunit superfamily / Urease, gamma subunit / Amidohydrolase family / Metal-dependent hydrolase, composite domain superfamily / Amidohydrolase-related / Metal-dependent hydrolase
Similarity search - Domain/homology
NICKEL (II) ION / HYDROXIDE ION / Urease subunit alpha / Urease subunit beta / Urease subunit gamma
Similarity search - Component
Biological speciesSporosarcina pasteurii (bacteria)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.12 Å
AuthorsMazzei, L. / Tria, G. / Ciurli, S. / Cianci, M.
Funding support1items
OrganizationGrant numberCountry
Not funded
CitationJournal: Int J Biol Macromol / Year: 2024
Title: Exploring the conformational space of the mobile flap in Sporosarcina pasteurii urease by cryo-electron microscopy.
Authors: Luca Mazzei / Giancarlo Tria / Stefano Ciurli / Michele Cianci /
Abstract: To fully understand enzymatic dynamics, it is essential to explore the complete conformational space of a biological catalyst. The catalytic mechanism of the nickel-dependent urease, the most ...To fully understand enzymatic dynamics, it is essential to explore the complete conformational space of a biological catalyst. The catalytic mechanism of the nickel-dependent urease, the most efficient enzyme known, holds significant relevance for medical, pharmaceutical, and agro-environmental applications. A critical aspect of urease function is the conformational change of a helix-turn-helix motif that covers the active site cavity, known as the mobile flap. This motif has been observed in either an open or a closed conformation through X-ray crystallography studies and has been proposed to stabilize the coordination of a urea molecule to the essential dinuclear Ni(II) cluster in the active site, a requisite for subsequent substrate hydrolysis. This study employs cryo-electron microscopy (cryo-EM) to investigate the transient states within the conformational space of the mobile flap, devoid of the possible constraints of crystallization conditions and solid-state effects. By comparing two cryo-EM structures of Sporosarcina pasteurii urease, one in its native form and the other inhibited by N-(n-butyl) phosphoric triamide (NBPTO), we have unprecedently identified an intermediate state between the open and the catalytically efficient closed conformation of the helix-turn-helix motif, suggesting a role of its tip region in this transition between the two states.
History
DepositionAug 29, 2024Deposition site: PDBE / Processing site: PDBE
Revision 1.0Jul 23, 2025Provider: repository / Type: Initial release

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: Urease subunit gamma
B: Urease subunit beta
C: Urease subunit alpha
hetero molecules


Theoretical massNumber of molelcules
Total (without water)86,8206
Polymers86,6863
Non-polymers1343
Water362
1


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

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Components

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Urease subunit ... , 3 types, 3 molecules ABC

#1: Protein Urease subunit gamma / Urea amidohydrolase subunit gamma


Mass: 11134.895 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Sporosarcina pasteurii (bacteria) / References: UniProt: P41022, urease
#2: Protein Urease subunit beta / Urea amidohydrolase subunit beta


Mass: 13975.539 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Sporosarcina pasteurii (bacteria) / References: UniProt: P41021, urease
#3: Protein Urease subunit alpha / Urea amidohydrolase subunit alpha


Mass: 61575.648 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Sporosarcina pasteurii (bacteria) / References: UniProt: P41020, urease

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Non-polymers , 3 types, 5 molecules

#4: Chemical ChemComp-NI / NICKEL (II) ION


Mass: 58.693 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: Ni
#5: Chemical ChemComp-OH / HYDROXIDE ION


Mass: 17.007 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: HO
#6: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Formula: H2O

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Details

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: native Sporosarcina pasteurii urease / Type: ORGANELLE OR CELLULAR COMPONENT / Entity ID: #1-#3 / Source: NATURAL
Molecular weightValue: 0.25 MDa / Experimental value: YES
Source (natural)Organism: Sporosarcina pasteurii (bacteria)
Buffer solutionpH: 7.5 / Details: 50 mM HEPES buffer, at pH 7.5
SpecimenConc.: 2 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: COPPER / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R1.2/1.3
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 283 K

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Electron microscopy imaging

MicroscopyModel: TFS GLACIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 200 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal defocus max: 2200 nm / Nominal defocus min: 800 nm / C2 aperture diameter: 70 µm
Image recordingAverage exposure time: 25 sec. / Electron dose: 30 e/Å2 / Detector mode: COUNTING / Film or detector model: FEI FALCON III (4k x 4k)

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Processing

EM softwareName: PHENIX / Category: model refinement
CTF correctionType: NONE
Particle selectionNum. of particles selected: 807538
SymmetryPoint symmetry: C3 (3 fold cyclic)
3D reconstructionResolution: 3.12 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 90461 / Symmetry type: POINT
Atomic model buildingProtocol: RIGID BODY FIT / Space: REAL
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.0036152
ELECTRON MICROSCOPYf_angle_d0.6558328
ELECTRON MICROSCOPYf_dihedral_angle_d5.242853
ELECTRON MICROSCOPYf_chiral_restr0.048941
ELECTRON MICROSCOPYf_plane_restr0.0041100

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