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- EMDB-34203: CryoEM structure of pentameric MotA from Aquifex aeolicus -

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Basic information

Entry
Database: EMDB / ID: EMD-34203
TitleCryoEM structure of pentameric MotA from Aquifex aeolicus
Map dataMotA 5mer postprocessed density map from dataset1
Sample
  • Complex: Pentameric MotA from Aquifex aeolicus
    • Protein or peptide: Motility protein A
Function / homology
Function and homology information


bacterial-type flagellum-dependent swarming motility / proton transmembrane transport / chemotaxis / membrane => GO:0016020 / plasma membrane
Similarity search - Function
Flagellar motor protein MotA, conserved site / Flagellar motor protein motA family signature. / MotA/TolQ/ExbB proton channel / MotA/TolQ/ExbB proton channel family
Similarity search - Domain/homology
Biological speciesAquifex aeolicus (bacteria) / Aquifex aeolicus VF5 (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.4 Å
AuthorsNishikino T / Takekawa N / Kishikawa J / Hirose M / Onoe S / Kato T / Imada K
Funding support Japan, 7 items
OrganizationGrant numberCountry
Japan Society for the Promotion of Science (JSPS)JP20J00329 Japan
Japan Society for the Promotion of Science (JSPS)JP16J01859 Japan
Japan Society for the Promotion of Science (JSPS)JP20K15732 Japan
Japan Society for the Promotion of Science (JSPS)JP20K06514 Japan
Japan Society for the Promotion of Science (JSPS)JP22K18359 Japan
Japan Society for the Promotion of Science (JSPS)JP22H02559 Japan
Japan Society for the Promotion of Science (JSPS)JP21H02443 Japan
CitationJournal: Biochem Biophys Res Commun / Year: 2022
Title: Structure of MotA, a flagellar stator protein, from hyperthermophile.
Authors: Tatsuro Nishikino / Norihiro Takekawa / Duy Phuoc Tran / Jun-Ichi Kishikawa / Mika Hirose / Sakura Onoe / Seiji Kojima / Michio Homma / Akio Kitao / Takayuki Kato / Katsumi Imada /
Abstract: Many motile bacteria swim and swarm toward favorable environments using the flagellum, which is rotated by a motor embedded in the inner membrane. The motor is composed of the rotor and the stator, ...Many motile bacteria swim and swarm toward favorable environments using the flagellum, which is rotated by a motor embedded in the inner membrane. The motor is composed of the rotor and the stator, and the motor torque is generated by the change of the interaction between the rotor and the stator induced by the ion flow through the stator. A stator unit consists of two types of membrane proteins termed A and B. Recent cryo-EM studies on the stators from mesophiles revealed that the stator consists of five A and two B subunits, whereas the low-resolution EM analysis showed that purified hyperthermophilic MotA forms a tetramer. To clarify the assembly formation and factors enhancing thermostability of the hyperthermophilic stator, we determined the cryo-EM structure of MotA from Aquifex aeolicus (Aa-MotA), a hyperthermophilic bacterium, at 3.42 Å resolution. Aa-MotA forms a pentamer with pseudo C5 symmetry. A simulated model of the Aa-MotAMotB stator complex resembles the structures of mesophilic stator complexes, suggesting that Aa-MotA can assemble into a pentamer equivalent to the stator complex without MotB. The distribution of hydrophobic residues of MotA pentamers suggests that the extremely hydrophobic nature in the subunit boundary and the transmembrane region is a key factor to stabilize hyperthermophilic Aa-MotA.
History
DepositionAug 31, 2022-
Header (metadata) releaseOct 26, 2022-
Map releaseOct 26, 2022-
UpdateNov 9, 2022-
Current statusNov 9, 2022Processing site: PDBj / Status: Released

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Structure visualization

Supplemental images

emd_34203.png

Downloads & links

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Map

FileDownload / File: emd_34203.map.gz / Format: CCP4 / Size: 103 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationMotA 5mer postprocessed density map from dataset1
Voxel sizeX=Y=Z: 0.9 Å
Density
Contour LevelBy AUTHOR: 0.02
Minimum - Maximum-0.10603084 - 0.16244212
Average (Standard dev.)4.768437e-05 (±0.004405161)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions300300300
Spacing300300300
CellA=B=C: 270.0 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_34203_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: half map 2 from dataset 1

Fileemd_34203_half_map_1.map
Annotationhalf map 2 from dataset 1
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: half map 1 from dataset 1

Fileemd_34203_half_map_2.map
Annotationhalf map 1 from dataset 1
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Pentameric MotA from Aquifex aeolicus

EntireName: Pentameric MotA from Aquifex aeolicus
Components
  • Complex: Pentameric MotA from Aquifex aeolicus
    • Protein or peptide: Motility protein A

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Supramolecule #1: Pentameric MotA from Aquifex aeolicus

SupramoleculeName: Pentameric MotA from Aquifex aeolicus / type: complex / Chimera: Yes / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Aquifex aeolicus (bacteria)
Recombinant expressionOrganism: Escherichia coli BL21 (bacteria)
Molecular weightTheoretical: 140 KDa

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Macromolecule #1: Motility protein A

MacromoleculeName: Motility protein A / type: protein_or_peptide / ID: 1
Details: Met1 to His6 is translation enhancing element sequence. 6 His residues on C-terminal are purification tag.
Number of copies: 5 / Enantiomer: LEVO
Source (natural)Organism: Aquifex aeolicus VF5 (bacteria) / Strain: VF5
Molecular weightTheoretical: 28.863195 KDa
Recombinant expressionOrganism: Escherichia coli BL21 (bacteria)
SequenceString: MNHKVHMDVG TIIGIIAAFL LILISILIGG SITAFINVPS IFIVVGGGMA AAMGAFPLKD FIRGVLAIKK AFLWKPPDLN DVIETIGEI ASKVRKEGIL ALEGDIELYY QKDPLLGDMI RMLVDGIDIN DIKATAEMAL AQLDEKMSTE VAVWEKLADL F PAFGMIGT ...String:
MNHKVHMDVG TIIGIIAAFL LILISILIGG SITAFINVPS IFIVVGGGMA AAMGAFPLKD FIRGVLAIKK AFLWKPPDLN DVIETIGEI ASKVRKEGIL ALEGDIELYY QKDPLLGDMI RMLVDGIDIN DIKATAEMAL AQLDEKMSTE VAVWEKLADL F PAFGMIGT LIGLIQMLRN LNDPSALGPG MAVALITTLY GAILANAFAI PVANKLKKAK DMEVLVKTIY IEAIEKIQKG EN PNVVKQE AAIMLGVELP EEVHHHHHH

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

BufferpH: 8
Component:
ConcentrationFormulaName
150.0 mMNaClSodium chloridesodium chloride
20.0 mMC4H11NO3Tris-HClTris
0.01 % (w/v)C47H88O22LMNG
GridModel: Quantifoil R1.2/1.3 / Material: MOLYBDENUM / Mesh: 300 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Atmosphere: AIR
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 60.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 1.8 µm / Nominal defocus min: 0.8 µm / Nominal magnification: 105000
Specialist opticsEnergy filter - Name: GIF Bioquantum / Energy filter - Slit width: 20 eV
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Number real images: 6299 / Average exposure time: 2.22 sec. / Average electron dose: 60.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 2240219
Startup modelType of model: PDB ENTRY
PDB model - PDB ID:

6ykm

Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.1)
Final 3D classificationNumber classes: 4 / Software - Name: RELION (ver. 3.1)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.1)
Final reconstructionNumber classes used: 1 / Applied symmetry - Point group: C1 (asymmetric) / Algorithm: FOURIER SPACE / Resolution.type: BY AUTHOR / Resolution: 3.4 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.1) / Number images used: 482752
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelPDB ID:

6ykm

RefinementSpace: REAL / Protocol: RIGID BODY FIT
Output model

PDB-8gqy:
CryoEM structure of pentameric MotA from Aquifex aeolicus

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