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- EMDB-21145: Structure of the bovine BBSome:ARL6:GTP complex -

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Basic information

Entry
Database: EMDB / ID: EMD-21145
TitleStructure of the bovine BBSome:ARL6:GTP complex
Map data
SampleBovine BBSome:ARL6:GTP complex
  • BBSome complex
  • (ADP-ribosylation factor-like protein ...) x 2
  • (Bardet-Biedl syndrome ...Bardet–Biedl syndrome) x 6
  • BBS1 domain-containing protein
  • Tetratricopeptide repeat domain 8
  • (ligand) x 2
Function / homology
Function and homology information


BBSome binding / protein transport from ciliary membrane to plasma membrane / protein localization to non-motile cilium / primary palate development / regulation of non-motile cilium assembly / multi-ciliated epithelial cell differentiation / negative regulation of appetite by leptin-mediated signaling pathway / renal tubule development / receptor localization to non-motile cilium / photoreceptor cell outer segment organization ...BBSome binding / protein transport from ciliary membrane to plasma membrane / protein localization to non-motile cilium / primary palate development / regulation of non-motile cilium assembly / multi-ciliated epithelial cell differentiation / negative regulation of appetite by leptin-mediated signaling pathway / renal tubule development / receptor localization to non-motile cilium / photoreceptor cell outer segment organization / axonemal microtubule / protein localization to photoreceptor outer segment / regulation of cilium beat frequency involved in ciliary motility / BBSome / camera-type eye photoreceptor cell differentiation / photoreceptor cell morphogenesis / retinal rod cell development / ventricular system development / ciliary transition zone / retina layer formation / establishment of planar polarity / smoothened binding / olfactory behavior / microtubule anchoring at centrosome / positive regulation of cilium assembly / olfactory bulb development / photoreceptor connecting cilium / negative regulation of systemic arterial blood pressure / inner ear receptor cell stereocilium organization / neural precursor cell proliferation / striatum development / patched binding / non-motile cilium / protein localization to cilium / cellular lipid metabolic process / non-motile cilium assembly / maintenance of protein location in nucleus / membrane coat / intracellular transport / hormone metabolic process / negative regulation of actin filament polymerization / brain morphogenesis / eye development / eating behavior / establishment of epithelial cell apical/basal polarity / positive regulation of multicellular organism growth / motile cilium / photoreceptor cell maintenance / centrosome cycle / limb development / phosphatidylinositol-3-phosphate binding / smoothened signaling pathway / regulation of smoothened signaling pathway / ciliary membrane / protein targeting to membrane / fertilization / sensory perception of smell / beta-tubulin binding / fat pad development / dynactin binding / fat cell differentiation / protein localization to centrosome / adult behavior / photoreceptor outer segment / cartilage development / spermatid development / regulation of stress fiber assembly / face development / ciliary basal body / negative regulation of GTPase activity / protein polymerization / pericentriolar material / alpha-tubulin binding / microtubule motor activity / centriolar satellite / cilium assembly / social behavior / dendrite development / RNA polymerase II repressing transcription factor binding / axoneme / mitotic cytokinesis / cilium / centriole / protein localization to plasma membrane / photoreceptor inner segment / microtubule organizing center / vesicle-mediated transport / cerebral cortex development / hippocampus development / protein localization / regulation of protein localization / regulation of cytokinesis / phospholipid binding / neuron migration / neural tube closure / phosphoprotein binding / intracellular protein transport / positive regulation of proteasomal ubiquitin-dependent protein catabolic process / microtubule cytoskeleton organization / protein transport
Tetratricopeptide-like helical domain superfamily / Bardet-Biedl syndrome 5 protein/sex-determination protein fem-3 / Parathyroid hormone-responsive B1 / P-loop containing nucleoside triphosphate hydrolase / PTHB1, N-terminal domain / Tetratricopeptide repeat-containing domain / Bardet-Biedl syndrome 7 protein / PTHB1, C-terminal domain / Cilia BBSome complex subunit 10 / Bardet-Biedl syndrome 1 protein ...Tetratricopeptide-like helical domain superfamily / Bardet-Biedl syndrome 5 protein/sex-determination protein fem-3 / Parathyroid hormone-responsive B1 / P-loop containing nucleoside triphosphate hydrolase / PTHB1, N-terminal domain / Tetratricopeptide repeat-containing domain / Bardet-Biedl syndrome 7 protein / PTHB1, C-terminal domain / Cilia BBSome complex subunit 10 / Bardet-Biedl syndrome 1 protein / DM16 repeat / Tetratricopeptide repeat protein 8 / Bardet-Biedl syndrome 2 protein / Ciliary BBSome complex subunit 2, middle region / Ciliary BBSome complex subunit 2, N-terminal / Ciliary BBSome complex subunit 2, C-terminal domain / Bardet-Biedl syndrome 1, N-terminal / ADP-ribosylation factor-like protein 6 / Quinoprotein alcohol dehydrogenase-like superfamily / Small GTPase superfamily, ARF/SAR type / Bardet-Biedl syndrome 5 protein / Small GTP-binding protein domain / Tetratricopeptide repeat / WD40-repeat-containing domain superfamily
Bardet-Biedl syndrome 5 protein homolog / Uncharacterized protein / BBS1 domain-containing protein / Bardet-Biedl syndrome 7 protein homolog / Tetratricopeptide repeat domain 8 / Uncharacterized protein / ADP-ribosylation factor-like protein 6 / Bardet-Biedl syndrome 4 protein homolog / Bardet-Biedl syndrome 2 protein homolog
Biological speciesBos taurus (cattle) / Bovine (cattle)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.5 Å
AuthorsSingh SK / Gui M / Koh F / Yip MCJ / Brown A
Funding support United States, 1 items
OrganizationGrant numberCountry
Other private United States
CitationJournal: Elife / Year: 2020
Title: Structure and activation mechanism of the BBSome membrane protein trafficking complex.
Authors: Sandeep K Singh / Miao Gui / Fujiet Koh / Matthew Cj Yip / Alan Brown /
Abstract: Bardet-Biedl syndrome (BBS) is a currently incurable ciliopathy caused by the failure to correctly establish or maintain cilia-dependent signaling pathways. Eight proteins associated with BBS ...Bardet-Biedl syndrome (BBS) is a currently incurable ciliopathy caused by the failure to correctly establish or maintain cilia-dependent signaling pathways. Eight proteins associated with BBS assemble into the BBSome, a key regulator of the ciliary membrane proteome. We report the electron cryomicroscopy (cryo-EM) structures of the native bovine BBSome in inactive and active states at 3.1 and 3.5 Å resolution, respectively. In the active state, the BBSome is bound to an Arf-family GTPase (ARL6/BBS3) that recruits the BBSome to ciliary membranes. ARL6 recognizes a composite binding site formed by BBS1 and BBS7 that is occluded in the inactive state. Activation requires an unexpected swiveling of the β-propeller domain of BBS1, the subunit most frequently implicated in substrate recognition, which widens a central cavity of the BBSome. Structural mapping of disease-causing mutations suggests that pathogenesis results from folding defects and the disruption of autoinhibition and activation.
Validation ReportPDB-ID: 6vbv

SummaryFull reportAbout validation report
History
DepositionDec 19, 2019-
Header (metadata) releaseJan 29, 2020-
Map releaseJan 29, 2020-
UpdateJan 29, 2020-
Current statusJan 29, 2020Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.02
  • Imaged by UCSF Chimera
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  • Surface view colored by radius
  • Surface level: 0.02
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-6vbv
  • Surface level: 0.02
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

Downloads & links

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Map

FileDownload / File: emd_21145.map.gz / Format: CCP4 / Size: 125 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.06 Å/pix.
x 320 pix.
= 339.2 Å
1.06 Å/pix.
x 320 pix.
= 339.2 Å
1.06 Å/pix.
x 320 pix.
= 339.2 Å

Surface

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Images are generated by Spider.

Voxel sizeX=Y=Z: 1.06 Å
Density
Contour LevelBy AUTHOR: 0.02 / Movie #1: 0.02
Minimum - Maximum0 - 1
Average (Standard dev.)0.030388517 (±0.16573176)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions320320320
Spacing320320320
CellA=B=C: 339.19998 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.061.061.06
M x/y/z320320320
origin x/y/z0.0000.0000.000
length x/y/z339.200339.200339.200
α/β/γ90.00090.00090.000
start NX/NY/NZ-38-19-20
NX/NY/NZ858082
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS320320320
D min/max/mean-0.0900.128-0.000

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Supplemental data

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Segmentation: #4

Fileemd_21145_msk_1.map
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Segmentation: #1

Fileemd_21145_msk_2.map
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Segmentation: #2

Fileemd_21145_msk_3.map
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Segmentation: #3

Fileemd_21145_msk_4.map
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Additional map: BBSome:ARL6:GTP complex. Unfiltered map

Fileemd_21145_additional_1.map
AnnotationBBSome:ARL6:GTP complex. Unfiltered map
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Additional map: BBSome:ARL6:GTP complex. Chimeric map following multibody refinement.

Fileemd_21145_additional_2.map
AnnotationBBSome:ARL6:GTP complex. Chimeric map following multibody refinement.
Projections & Slices
AxesZYX

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Additional map: Post-processed multibody map for the BBSome body

Fileemd_21145_additional_3.map
AnnotationPost-processed multibody map for the BBSome body
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Additional map: Post-processed multibody map for the BBSome head

Fileemd_21145_additional_4.map
AnnotationPost-processed multibody map for the BBSome head
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Additional map: Post-processed multibody map for the BBS1-ARL6 interaction.

Fileemd_21145_additional_5.map
AnnotationPost-processed multibody map for the BBS1-ARL6 interaction.
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Half map: BBSome:ARL6:GTP complex. Half map 1

Fileemd_21145_half_map_1.map
AnnotationBBSome:ARL6:GTP complex. Half map 1
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Half map: BBSome:ARL6:GTP complex. Half map 2

Fileemd_21145_half_map_2.map
AnnotationBBSome:ARL6:GTP complex. Half map 2
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Sample components

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Entire Bovine BBSome:ARL6:GTP complex

EntireName: Bovine BBSome:ARL6:GTP complex
Details: Native BBSome complex isolated from bovine retina and incubated with recombinant bovine ARL6 in the presence of GTP.
Number of components: 14

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Component #1: protein, Bovine BBSome:ARL6:GTP complex

ProteinName: Bovine BBSome:ARL6:GTP complex
Details: Native BBSome complex isolated from bovine retina and incubated with recombinant bovine ARL6 in the presence of GTP.
Recombinant expression: No
MassTheoretical: 500 kDa

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Component #2: protein, BBSome complex

ProteinName: BBSome complex / Recombinant expression: No
SourceSpecies: Bos taurus (cattle)
Source (natural)Organ or tissue: Eye

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Component #3: protein, ADP-ribosylation factor-like protein 6

ProteinName: ADP-ribosylation factor-like protein 6 / Recombinant expression: No
SourceSpecies: Bos taurus (cattle)
Source (engineered)Expression System: Escherichia coli BL21(DE3) (bacteria) / Strain: BL21(DE3)

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Component #4: protein, Bardet-Biedl syndrome 18 protein

ProteinName: Bardet-Biedl syndrome 18 proteinBardet–Biedl syndrome
Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 8.070502 kDa
SourceSpecies: Bovine (cattle)
Source (natural)Organ or tissue: Eye

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Component #5: protein, BBS1 domain-containing protein

ProteinName: BBS1 domain-containing protein / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 64.939141 kDa
SourceSpecies: Bovine (cattle)
Source (natural)Organ or tissue: Eye

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Component #6: protein, Bardet-Biedl syndrome 2 protein homolog

ProteinName: Bardet-Biedl syndrome 2 protein homologBardet–Biedl syndrome
Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 79.911484 kDa
SourceSpecies: Bovine (cattle)
Source (natural)Organ or tissue: Eye

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Component #7: protein, Bardet-Biedl syndrome 4 protein homolog

ProteinName: Bardet-Biedl syndrome 4 protein homologBardet–Biedl syndrome
Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 58.289133 kDa
SourceSpecies: Bovine (cattle)
Source (natural)Organ or tissue: Eye

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Component #8: protein, Bardet-Biedl syndrome 5 protein homolog

ProteinName: Bardet-Biedl syndrome 5 protein homologBardet–Biedl syndrome
Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 38.880984 kDa
SourceSpecies: Bovine (cattle)
Source (natural)Organ or tissue: Eye

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Component #9: protein, Bardet-Biedl syndrome 7 protein homolog

ProteinName: Bardet-Biedl syndrome 7 protein homologBardet–Biedl syndrome
Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 80.471375 kDa
SourceSpecies: Bovine (cattle)
Source (natural)Organ or tissue: Eye

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Component #10: protein, Tetratricopeptide repeat domain 8

ProteinName: Tetratricopeptide repeat domain 8 / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 56.686406 kDa
SourceSpecies: Bovine (cattle)
Source (natural)Organ or tissue: Eye

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Component #11: protein, Bardet-Biedl syndrome 9

ProteinName: Bardet-Biedl syndrome 9Bardet–Biedl syndrome / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 99.230914 kDa
SourceSpecies: Bovine (cattle)
Source (natural)Organ or tissue: Eye

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Component #12: protein, ADP-ribosylation factor-like protein 6

ProteinName: ADP-ribosylation factor-like protein 6 / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 21.086486 kDa
SourceSpecies: Bos taurus (cattle)
Source (engineered)Expression System: Escherichia coli BL21(DE3) (bacteria)

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Component #13: ligand, CALCIUM ION

LigandName: CALCIUM IONCalcium / Number of Copies: 2 / Recombinant expression: No
MassTheoretical: 4.007805 MDa

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Component #14: ligand, GUANOSINE-5'-TRIPHOSPHATE

LigandName: GUANOSINE-5'-TRIPHOSPHATEGuanosine triphosphate / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 0.52318 kDa

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Experimental details

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Sample preparation

SpecimenSpecimen state: Particle / Method: cryo EM
Sample solutionSpecimen conc.: 0.7 mg/mL / pH: 7.5
Support filmunspecified
VitrificationInstrument: FEI VITROBOT MARK II / Cryogen name: ETHANE / Temperature: 293 K / Humidity: 100 % / Details: Grids were blotted for 2 s with a -2 offset..

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
ImagingMicroscope: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Electron dose: 56 e/Å2 / Illumination mode: FLOOD BEAM
LensMagnification: 81000.0 X (nominal) / Cs: 2.7 mm / Imaging mode: BRIGHT FIELD / Defocus: 1100.0 - 2400.0 nm / Energy filter: GIF Bioquantum
Specimen HolderModel: FEI TITAN KRIOS AUTOGRID HOLDER
CameraDetector: OTHER

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Image acquisition

Image acquisitionNumber of digital images: 9408

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Image processing

ProcessingMethod: single particle reconstruction / Applied symmetry: C1 (asymmetric) / Number of projections: 75201
3D reconstructionAlgorithm: BACK PROJECTION / Software: RELION / Resolution: 3.5 Å / Resolution method: FSC 0.143 CUT-OFF
FSC plot (resolution estimation)

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Atomic model buiding

Modeling #1Target criteria: Correlation coefficient / Refinement space: REAL
Details: During refinement, the resolution limit was set to 3.5 Angstrom. Secondary structure, Ramachandran and rotamer restraints were applied during refinement.
Overall bvalue: 43.6
Output model

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