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- PDB-9ijm: Bacterial flagellar sodium-driven stator PomA5PomB2 with 100 mM N... -

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Basic information

Entry
Database: PDB / ID: 9ijm
TitleBacterial flagellar sodium-driven stator PomA5PomB2 with 100 mM NaCl and 0.1 mM phenamil
Components
  • Chemotaxis protein PomA
  • PomB
KeywordsMEMBRANE PROTEIN / bacterial flagellar stator unit / Phenamil / sodium ion transport / membrane protein complex
Function / homology
Function and homology information


bacterial-type flagellum-dependent swarming motility / proton transmembrane transport / chemotaxis / plasma membrane
Similarity search - Function
Flagellar motor protein MotA, conserved site / Flagellar motor protein motA family signature. / : / Motility protein B-like, N-terminal domain / Membrane MotB of proton-channel complex MotA/MotB / MotA/TolQ/ExbB proton channel / MotA/TolQ/ExbB proton channel family / : / OmpA-like domain profile. / OmpA-like domain superfamily ...Flagellar motor protein MotA, conserved site / Flagellar motor protein motA family signature. / : / Motility protein B-like, N-terminal domain / Membrane MotB of proton-channel complex MotA/MotB / MotA/TolQ/ExbB proton channel / MotA/TolQ/ExbB proton channel family / : / OmpA-like domain profile. / OmpA-like domain superfamily / OmpA family / OmpA-like domain
Similarity search - Domain/homology
: / Chemotaxis protein PomA / PomB
Similarity search - Component
Biological speciesVibrio alginolyticus (bacteria)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.32 Å
AuthorsNishikino, T. / Takekawa, N. / Imada, K.
Funding support Japan, 2items
OrganizationGrant numberCountry
Japan Society for the Promotion of Science (JSPS)JP20J00329 Japan
Japan Society for the Promotion of Science (JSPS)JP23K14157 Japan
Citation
Journal: Proc Natl Acad Sci U S A / Year: 2025
Title: Structural insight into sodium ion pathway in the bacterial flagellar stator from marine .
Authors: Tatsuro Nishikino / Norihiro Takekawa / Jun-Ichi Kishikawa / Mika Hirose / Seiji Kojima / Michio Homma / Takayuki Kato / Katsumi Imada /
Abstract: Many bacteria swim in liquid or swarm on surface using the flagellum rotated by a motor driven by specific ion flow. The motor consists of the rotor and stator, and the stator converts the energy of ...Many bacteria swim in liquid or swarm on surface using the flagellum rotated by a motor driven by specific ion flow. The motor consists of the rotor and stator, and the stator converts the energy of ion flow to mechanical rotation. However, the ion pathway and the mechanism of stator rotation coupled with specific ion flow are still obscure. Here, we determined the structures of the sodium-driven stator of , namely PomAB, in the presence and absence of sodium ions and the structure with its specific inhibitor, phenamil, by cryo-electron microscopy. The structures and following functional analysis revealed the sodium ion pathway, the mechanism of ion selectivity, and the inhibition mechanism by phenamil. We propose a model of sodium ion flow coupled with the stator rotation based on the structures. This work provides insights into the molecular mechanisms of ion specificity and conversion of the electrochemical potential into mechanical functions.
#1: Journal: Proc.Natl.Acad.Sci.USA
Title: Structural insight into sodium ion pathway in the bacterial flagellar stator from marine Vibrio
Authors: Nishikino, T. / Takekawa, N. / Kishikawa, J. / Hirose, M. / Kojima, S. / Homma, M. / Kato, T. / Imada, K.
History
DepositionJun 24, 2024Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0Jan 1, 2025Provider: repository / Type: Initial release
Revision 1.1Jan 22, 2025Group: Data collection / Database references / Category: citation / citation_author / em_admin / Item: _em_admin.last_update

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: PomB
B: PomB
C: Chemotaxis protein PomA
D: Chemotaxis protein PomA
E: Chemotaxis protein PomA
F: Chemotaxis protein PomA
G: Chemotaxis protein PomA
hetero molecules


Theoretical massNumber of molelcules
Total (without water)209,42710
Polymers209,0757
Non-polymers3523
Water181
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: gel filtration
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: Protein PomB


Mass: 36330.035 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Details: 6 His residues on C-terminal are purification tag / Source: (gene. exp.) Vibrio alginolyticus (bacteria) / Strain: VIO5 / Gene: pomB / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: O06874
#2: Protein
Chemotaxis protein PomA


Mass: 27283.031 Da / Num. of mol.: 5
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Vibrio alginolyticus (bacteria) / Strain: VIO5 / Gene: pomA / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: O06873
#3: Chemical ChemComp-NA / SODIUM ION


Mass: 22.990 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: Na / Feature type: SUBJECT OF INVESTIGATION
#4: Chemical ChemComp-A1L2K / phenamil / 3,5-bis(azanyl)-6-chloranyl-~{N}-(~{N}-phenylcarbamimidoyl)pyrazine-2-carboxamide


Mass: 305.723 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C12H12ClN7O / Feature type: SUBJECT OF INVESTIGATION
#5: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Formula: H2O
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: Bacterial flagellar sodium-driven stator PomA5PomB2 with 100 mM NaCl and 0.1 mM phenamil
Type: COMPLEX / Entity ID: #1-#2 / Source: RECOMBINANT
Molecular weightValue: 0.21 MDa / Experimental value: YES
Source (natural)Organism: Vibrio alginolyticus (bacteria) / Strain: VIO5
Source (recombinant)Organism: Escherichia coli BL21(DE3) (bacteria)
Buffer solutionpH: 8
Buffer component
IDConc.NameFormulaBuffer-ID
1100 mMsodium chlorideNaCl1
220 mMTris-HClC4H11NO31
30.005 % (w/v)GDNC56H92O251
41 % (w/v)DMSOC2H6SO1
50.1 mMPhenamil methanesulfonate saltC12H12ClN7O/CH3SO3H1
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: COPPER / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R0.6/1
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277 K

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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: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal magnification: 105000 X / Nominal defocus max: 2800 nm / Nominal defocus min: 725 nm / C2 aperture diameter: 50 µm / Alignment procedure: COMA FREE
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingElectron dose: 60 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Num. of real images: 13874
EM imaging opticsEnergyfilter name: GIF Bioquantum / Energyfilter slit width: 20 eV

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Processing

EM software
IDNameVersionCategory
1RELION4particle selection
2SerialEMimage acquisition
4CTFFIND4.1CTF correction
7Coot0.9.8.7model fitting
9RELION4initial Euler assignment
10RELION4final Euler assignment
11RELION4classification
12RELION43D reconstruction
13PHENIXmodel refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 4573172
3D reconstructionResolution: 3.32 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 596902 / Algorithm: FOURIER SPACE / Symmetry type: POINT
Atomic model buildingProtocol: RIGID BODY FIT / Space: REAL
Atomic model buildingPDB-ID: 8ZYV

8zyv


Accession code: 8ZYV / Source name: PDB / Type: experimental model
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.0068833
ELECTRON MICROSCOPYf_angle_d0.66111899
ELECTRON MICROSCOPYf_dihedral_angle_d4.7481194
ELECTRON MICROSCOPYf_chiral_restr0.0431409
ELECTRON MICROSCOPYf_plane_restr0.0041511

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