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- EMDB-4992: Escherichia coli chemotaxis signaling arrays at high kinase activ... -

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Entry
Database: EMDB / ID: EMD-4992
TitleEscherichia coli chemotaxis signaling arrays at high kinase activity with serine receptor mutant Tsr_QQQQ
Map dataEscherichia coli chemotaxis signaling arrays at high kinase activity with serine receptor mutant Tsr_QQQQ
Sample
  • Cell: Escherichia coli chemotaxis signaling arrays at high kinase activity with serine receptor mutant Tsr_QQQQ
Biological speciesEscherichia coli (E. coli)
Methodsubtomogram averaging / cryo EM / Resolution: 20.1 Å
AuthorsYang W / Cassidy CK / Ames P / Diebolder CA / Schulten K / Luthey-Schulten Z / Parkinson JS / Briegel A
CitationJournal: mBio / Year: 2019
Title: Conformational Changes of the Escherichia coli Serine Chemoreceptor in Different Signaling States.
Authors: Wen Yang / C Keith Cassidy / Peter Ames / Christoph A Diebolder / Klaus Schulten / Zaida Luthey-Schulten / John S Parkinson / Ariane Briegel /
Abstract: Tsr, the serine chemoreceptor in , transduces signals from a periplasmic ligand-binding site to its cytoplasmic tip, where it controls the activity of the CheA kinase. To function, Tsr forms trimers ...Tsr, the serine chemoreceptor in , transduces signals from a periplasmic ligand-binding site to its cytoplasmic tip, where it controls the activity of the CheA kinase. To function, Tsr forms trimers of homodimers (TODs), which associate with the CheA kinase and CheW coupling protein. Together, these proteins assemble into extended hexagonal arrays. Here, we use cryo-electron tomography and molecular dynamics simulation to study Tsr in the context of a near-native array, characterizing its signaling-related conformational changes at both the individual dimer and the trimer level. In particular, we show that individual Tsr dimers within a trimer exhibit asymmetric flexibilities that are a function of the signaling state, highlighting the effect of their different protein interactions at the receptor tips. We further reveal that the dimer compactness of the Tsr trimer changes between signaling states, transitioning at the glycine hinge from a compact conformation in the kinase-OFF state to an expanded conformation in the kinase-ON state. Hence, our results support a crucial role for the glycine hinge: to allow the receptor flexibility necessary to achieve different signaling states while also maintaining structural constraints imposed by the membrane and extended array architecture. In , membrane-bound chemoreceptors, the histidine kinase CheA, and coupling protein CheW form highly ordered chemosensory arrays. In core signaling complexes, chemoreceptor trimers of dimers undergo conformational changes, induced by ligand binding and sensory adaptation, which regulate kinase activation. Here, we characterize by cryo-electron tomography the kinase-ON and kinase-OFF conformations of the serine receptor in its native array context. We found distinctive structural differences between the members of a receptor trimer, which contact different partners in the signaling unit, and structural differences between the ON and OFF signaling complexes. Our results provide new insights into the signaling mechanism of chemoreceptor arrays and suggest an important functional role for a previously postulated flexible region and glycine hinge in the receptor molecule.
History
DepositionMay 22, 2019-
Header (metadata) releaseJun 12, 2019-
Map releaseJul 17, 2019-
UpdateJul 17, 2019-
Current statusJul 17, 2019Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 1.05
  • Imaged by UCSF Chimera
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  • Surface view colored by height
  • Surface level: 1.05
  • Imaged by UCSF Chimera
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Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

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Map

FileDownload / File: emd_4992.map.gz / Format: CCP4 / Size: 3 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationEscherichia coli chemotaxis signaling arrays at high kinase activity with serine receptor mutant Tsr_QQQQ
Voxel sizeX=Y=Z: 3.513 Å
Density
Contour LevelBy AUTHOR: 1.05 / Movie #1: 1.05
Minimum - Maximum-3.2930236 - 3.5379412
Average (Standard dev.)-0.005995805 (±0.71703994)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin-46-46-46
Dimensions929292
Spacing929292
CellA=B=C: 323.196 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z3.5133.5133.513
M x/y/z929292
origin x/y/z0.0000.0000.000
length x/y/z323.196323.196323.196
α/β/γ90.00090.00090.000
start NX/NY/NZ000
NX/NY/NZ300300300
MAP C/R/S123
start NC/NR/NS-46-46-46
NC/NR/NS929292
D min/max/mean-3.2933.538-0.006

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Supplemental data

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Sample components

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Entire : Escherichia coli chemotaxis signaling arrays at high kinase activ...

EntireName: Escherichia coli chemotaxis signaling arrays at high kinase activity with serine receptor mutant Tsr_QQQQ
Components
  • Cell: Escherichia coli chemotaxis signaling arrays at high kinase activity with serine receptor mutant Tsr_QQQQ

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Supramolecule #1: Escherichia coli chemotaxis signaling arrays at high kinase activ...

SupramoleculeName: Escherichia coli chemotaxis signaling arrays at high kinase activity with serine receptor mutant Tsr_QQQQ
type: cell / ID: 1 / Parent: 0
Source (natural)Organism: Escherichia coli (E. coli)

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Experimental details

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Structure determination

Methodcryo EM
Processingsubtomogram averaging
Aggregation statecell

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Sample preparation

BufferpH: 7.5
GridModel: Quantifoil R2/2 / Material: COPPER / Mesh: 200
VitrificationCryogen name: ETHANE

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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
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Average electron dose: 1.3 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

ExtractionNumber tomograms: 11 / Number images used: 3504 / Software - Name: Dynamo
Final angle assignmentType: NOT APPLICABLE / Software - Name: CTFPHASEFLIP
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 20.1 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: Dynamo / Number subtomograms used: 1977

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