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- EMDB-30379: 11-fold symmetry of Salmonella FliF-FliG ring complex -

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Basic information

Entry
Database: EMDB / ID: EMD-30379
Title11-fold symmetry of Salmonella FliF-FliG ring complex
Map data
Sample
  • Complex: 11-fold symmetry of Salmonella FliF-FliG ring complex
Function / homology
Function and homology information


bacterial-type flagellum basal body, MS ring / cytoskeletal motor activity / bacterial-type flagellum-dependent cell motility / membrane => GO:0016020 / plasma membrane
Similarity search - Function
Flagellar M-ring protein FliF / Flagellar M-ring C-terminal / Flagellar M-ring protein C-terminal / Lipoprotein YscJ/Flagellar M-ring protein / Secretory protein of YscJ/FliF family / Flagellar M-ring , N-terminal
Similarity search - Domain/homology
Flagellar M-ring protein
Similarity search - Component
Biological speciesSalmonella enterica subsp. enterica serovar Typhimurium (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 6.1 Å
AuthorsKawamoto A / Kato T / Kinoshita M / Minamino T / Namba K
Funding support Japan, 10 items
OrganizationGrant numberCountry
Japan Society for the Promotion of Science (JSPS)JP16J01859 Japan
Japan Society for the Promotion of Science (JSPS)JP18K14639 Japan
Japan Society for the Promotion of Science (JSPS)JP18K06155 Japan
Japan Society for the Promotion of Science (JSPS)JP18K06155 Japan
Japan Society for the Promotion of Science (JSPS)JP18K14638 Japan
Japan Society for the Promotion of Science (JSPS)JP19H03182 Japan
Japan Society for the Promotion of Science (JSPS)JP25000013 Japan
Japan Society for the Promotion of Science (JSPS)JP15H02386 Japan
Japan Agency for Medical Research and Development (AMED)JP19am0101117 Japan
Japan Agency for Medical Research and Development (AMED)JP17pc0101020 Japan
CitationJournal: mBio / Year: 2021
Title: Two Distinct Conformations in 34 FliF Subunits Generate Three Different Symmetries within the Flagellar MS-Ring.
Authors: Norihiro Takekawa / Akihiro Kawamoto / Mayuko Sakuma / Takayuki Kato / Seiji Kojima / Miki Kinoshita / Tohru Minamino / Keiichi Namba / Michio Homma / Katsumi Imada /
Abstract: The bacterial flagellum is a protein nanomachine essential for bacterial motility. The flagellar basal body contains several ring structures. The MS-ring is embedded in the cytoplasmic membrane and ...The bacterial flagellum is a protein nanomachine essential for bacterial motility. The flagellar basal body contains several ring structures. The MS-ring is embedded in the cytoplasmic membrane and is formed at the earliest stage of flagellar formation to serve as the base for flagellar assembly as well as a housing for the flagellar protein export gate complex. The MS-ring is formed by FliF, which has two transmembrane helices and a large periplasmic region. A recent electron cryomicroscopy (cryoEM) study of the MS-ring formed by overexpressed FliF revealed a symmetry mismatch between the S-ring and inner part of the M-ring. However, the actual symmetry relation in the native MS-ring and positions of missing domains remain obscure. Here, we show the structure of the M-ring by combining cryoEM and X-ray crystallography. The crystal structure of the N-terminal half of the periplasmic region of FliF showed that it consists of two domains (D1 and D2) resembling PrgK D1/PrgH D2 and PrgK D2/PrgH D3 of the injectisome. CryoEM analysis revealed that the inner part of the M-ring shows a gear wheel-like density with the inner ring of C23 symmetry surrounded by cogs with C11 symmetry, to which 34 copies of FliF fitted well. We propose that FliF adopts two distinct orientations in the M-ring relative to the rest of FliF, with 23 chains forming the wheel and 11 chains forming the cogs, and the 34 chains come together to form the S-ring with C34 symmetry for multiple functions of the MS-ring. The bacterial flagellum is a motility organelle formed by tens of thousands of protein molecules. At the earliest stage of flagellar assembly, a transmembrane protein, FliF, forms the MS-ring in the cytoplasmic membrane as the base for flagellar assembly. Here, we solved the crystal structure of a FliF fragment. Electron cryomicroscopy (cryoEM) structural analysis of the MS-ring showed that the M-ring and S-ring have different rotational symmetries. By docking the crystal structure of the FliF fragment into the cryoEM density map of the entire MS-ring, we built a model of the whole periplasmic region of FliF and proposed that FliF adopts two distinct conformations to generate three distinct C11, C23, and C34 symmetries within the MS-ring for its multiple functions.
History
DepositionJul 14, 2020-
Header (metadata) releaseFeb 17, 2021-
Map releaseFeb 17, 2021-
UpdateSep 1, 2021-
Current statusSep 1, 2021Processing site: PDBj / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.0158
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by cylindrical radius
  • Surface level: 0.0158
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_30379.png

Downloads & links

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Map

FileDownload / File: emd_30379.map.gz / Format: CCP4 / Size: 421.9 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Voxel sizeX=Y=Z: 1.07 Å
Density
Contour LevelBy AUTHOR: 0.0158 / Movie #1: 0.0158
Minimum - Maximum-0.034747247 - 0.10156517
Average (Standard dev.)0.0006567562 (±0.0044418843)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions480480480
Spacing480480480
CellA=B=C: 513.60004 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.071.071.07
M x/y/z480480480
origin x/y/z0.0000.0000.000
length x/y/z513.600513.600513.600
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS480480480
D min/max/mean-0.0350.1020.001

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Supplemental data

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Sample components

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Entire : 11-fold symmetry of Salmonella FliF-FliG ring complex

EntireName: 11-fold symmetry of Salmonella FliF-FliG ring complex
Components
  • Complex: 11-fold symmetry of Salmonella FliF-FliG ring complex

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Supramolecule #1: 11-fold symmetry of Salmonella FliF-FliG ring complex

SupramoleculeName: 11-fold symmetry of Salmonella FliF-FliG ring complex / type: complex / ID: 1 / Parent: 0
Source (natural)Organism: Salmonella enterica subsp. enterica serovar Typhimurium (bacteria)

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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:
ConcentrationFormula
25.0 mMTris-HClTris
50.0 mMNaClSodium chloride
1.0 mMEDTAEthylenediaminetetraacetic acid
0.1 %DDM
GridModel: Quantifoil R0.6/1 / Material: COPPER / Mesh: 200 / Pretreatment - Type: GLOW DISCHARGE
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Instrument: FEI VITROBOT MARK III

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 3.0 µm / Nominal defocus min: 1.0 µm / Nominal magnification: 75000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Image recordingFilm or detector model: FEI FALCON II (4k x 4k) / Number real images: 6961 / Average electron dose: 90.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 6961
CTF correctionSoftware - Name: Gctf
Startup modelType of model: OTHER
Details: the initial 3D model of the MS-ring using cryoSPARC
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 2.1)
Final 3D classificationSoftware - Name: RELION (ver. 2.1)
Final angle assignmentType: ANGULAR RECONSTITUTION / Software - Name: RELION (ver. 2.1)
Final reconstructionApplied symmetry - Point group: C11 (11 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 6.1 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 2.1) / Number images used: 53522

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