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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 dataBBSome:ARL6:GTP complex. Post-processed map.
Sample
  • Complex: Bovine BBSome:ARL6:GTP complex
    • Complex: BBSome complex
      • Protein or peptide: x 8 types
    • Complex: ADP-ribosylation factor-like protein 6
      • Protein or peptide: x 1 types
  • Ligand: x 2 types
KeywordsCilia / ciliopathy / complex / membrane-protein transport / PROTEIN TRANSPORT
Function / homology
Function and homology information


BBSome binding / establishment of anatomical structure orientation / BBSome-mediated cargo-targeting to cilium / multi-ciliated epithelial cell differentiation / receptor localization to non-motile cilium / renal tubule development / BBSome / camera-type eye photoreceptor cell differentiation / smoothened binding / establishment of planar polarity ...BBSome binding / establishment of anatomical structure orientation / BBSome-mediated cargo-targeting to cilium / multi-ciliated epithelial cell differentiation / receptor localization to non-motile cilium / renal tubule development / BBSome / camera-type eye photoreceptor cell differentiation / smoothened binding / establishment of planar polarity / axonemal microtubule / photoreceptor connecting cilium / inner ear receptor cell stereocilium organization / patched binding / olfactory bulb development / membrane coat / protein localization to cilium / non-motile cilium assembly / phosphatidylinositol-3-phosphate binding / establishment of epithelial cell apical/basal polarity / non-motile cilium / regulation of stress fiber assembly / motile cilium / protein targeting to membrane / centrosome cycle / ciliary membrane / pericentriolar material / fat cell differentiation / axoneme / intracellular transport / cilium assembly / centriolar satellite / protein polymerization / vesicle-mediated transport / ciliary basal body / axon guidance / protein localization to plasma membrane / phospholipid binding / intracellular protein transport / multicellular organism growth / protein localization / cilium / regulation of protein localization / sensory perception of smell / protein-macromolecule adaptor activity / protein transport / RNA polymerase II-specific DNA-binding transcription factor binding / neuron projection / GTPase activity / centrosome / GTP binding / membrane / cytoplasm
Similarity search - Function
ADP-ribosylation factor-like protein 6 / Bardet-Biedl syndrome 7 protein / Bardet-Biedl syndrome 2 protein / Ciliary BBSome complex subunit 2, C-terminal domain / Ciliary BBSome complex subunit 2, middle region / Ciliary BBSome complex subunit 2, N-terminal / Ciliary BBSome complex subunit 2, N-terminal / Ciliary BBSome complex subunit 2, C-terminal / Ciliary BBSome complex subunit 2, middle region / Bardet-Biedl syndrome 5 protein ...ADP-ribosylation factor-like protein 6 / Bardet-Biedl syndrome 7 protein / Bardet-Biedl syndrome 2 protein / Ciliary BBSome complex subunit 2, C-terminal domain / Ciliary BBSome complex subunit 2, middle region / Ciliary BBSome complex subunit 2, N-terminal / Ciliary BBSome complex subunit 2, N-terminal / Ciliary BBSome complex subunit 2, C-terminal / Ciliary BBSome complex subunit 2, middle region / Bardet-Biedl syndrome 5 protein / DM16 repeat / Cilia BBSome complex subunit 10 / Tetratricopeptide repeat protein 8 / Bardet-Biedl syndrome 5 protein/sex-determination protein fem-3 / Bardet-Biedl syndrome 5 protein / Cilia BBSome complex subunit 10 / Repeats in sea squirt COS41.4, worm R01H10.6, fly CG1126 etc. / Parathyroid hormone-responsive B1 / PTHB1, N-terminal domain / PTHB1, C-terminal domain / PTHB1 N-terminus / PTHB1 C-terminus / Bardet-Biedl syndrome 1 protein / Bardet-Biedl syndrome 1, N-terminal / Ciliary BBSome complex subunit 1 / TPR repeat / Tetratricopeptide repeat / small GTPase Arf family profile. / Sar1p-like members of the Ras-family of small GTPases / Tetratricopeptide repeat / Small GTPase superfamily, ARF/SAR type / ADP-ribosylation factor family / ARF-like small GTPases; ARF, ADP-ribosylation factor / Tetratricopeptide repeat / Quinoprotein alcohol dehydrogenase-like superfamily / TPR repeat region circular profile. / TPR repeat profile. / Tetratricopeptide repeats / Tetratricopeptide repeat / Tetratricopeptide-like helical domain superfamily / Small GTP-binding protein domain / PH-like domain superfamily / WD40-repeat-containing domain superfamily / WD40/YVTN repeat-like-containing domain superfamily / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
Bardet-Biedl syndrome 5 protein homolog / Bardet-Biedl syndrome 9 / Bardet-Biedl syndrome 1 / Bardet-Biedl syndrome 7 protein homolog / Tetratricopeptide repeat domain 8 / BBSome interacting protein 1 / ADP-ribosylation factor-like protein 6 / Bardet-Biedl syndrome 4 protein homolog / Bardet-Biedl syndrome 2 protein homolog
Similarity search - Component
Biological speciesBos taurus (cattle)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.5 Å
AuthorsSingh SK / Gui M
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.
History
DepositionDec 19, 2019-
Header (metadata) releaseJan 29, 2020-
Map releaseJan 29, 2020-
UpdateMar 6, 2024-
Current statusMar 6, 2024Processing 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
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_21145.png

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)
AnnotationBBSome:ARL6:GTP complex. Post-processed map.
Voxel sizeX=Y=Z: 1.06 Å
Density
Contour LevelBy AUTHOR: 0.02 / Movie #1: 0.02
Minimum - Maximum-0.09028247 - 0.1276134
Average (Standard dev.)-0.00007061716 (±0.004018769)
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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Mask #1

Fileemd_21145_msk_1.map
Projections & Slices
AxesZYX

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Mask #2

Fileemd_21145_msk_2.map
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Mask #3

Fileemd_21145_msk_3.map
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Mask #4

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
Projections & Slices
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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
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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
Components
  • Complex: Bovine BBSome:ARL6:GTP complex
    • Complex: BBSome complex
      • Protein or peptide: Bardet-Biedl syndrome 18 proteinBardet–Biedl syndrome
      • Protein or peptide: BBS1 domain-containing protein
      • Protein or peptide: Bardet-Biedl syndrome 2 protein homologBardet–Biedl syndrome
      • Protein or peptide: Bardet-Biedl syndrome 4 protein homologBardet–Biedl syndrome
      • Protein or peptide: Bardet-Biedl syndrome 5 protein homologBardet–Biedl syndrome
      • Protein or peptide: Bardet-Biedl syndrome 7 protein homologBardet–Biedl syndrome
      • Protein or peptide: Tetratricopeptide repeat domain 8Tetratricopeptide repeat
      • Protein or peptide: Bardet-Biedl syndrome 9Bardet–Biedl syndrome
    • Complex: ADP-ribosylation factor-like protein 6
      • Protein or peptide: ADP-ribosylation factor-like protein 6
  • Ligand: CALCIUM IONCalcium
  • Ligand: GUANOSINE-5'-TRIPHOSPHATEGuanosine triphosphate

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Supramolecule #1: Bovine BBSome:ARL6:GTP complex

SupramoleculeName: Bovine BBSome:ARL6:GTP complex / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#9
Details: Native BBSome complex isolated from bovine retina and incubated with recombinant bovine ARL6 in the presence of GTP.
Molecular weightTheoretical: 500 KDa

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Supramolecule #2: BBSome complex

SupramoleculeName: BBSome complex / type: complex / ID: 2 / Parent: 1 / Macromolecule list: #1-#8
Source (natural)Organism: Bos taurus (cattle) / Organ: Eye / Tissue: Retina

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Supramolecule #3: ADP-ribosylation factor-like protein 6

SupramoleculeName: ADP-ribosylation factor-like protein 6 / type: complex / ID: 3 / Parent: 1 / Macromolecule list: #9
Source (natural)Organism: Bos taurus (cattle)

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Macromolecule #1: Bardet-Biedl syndrome 18 protein

MacromoleculeName: Bardet-Biedl syndrome 18 protein / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Bos taurus (cattle) / Organ: Eye / Tissue: Retina
Molecular weightTheoretical: 8.070502 KDa
SequenceString:
MAETKSMFRE VLPKQGQLYV EDITTMVLCK PKLLPLKSLT LEKLEKMQQA AQDTIHQQEM TEKEQKITH

UniProtKB: BBSome interacting protein 1

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Macromolecule #2: BBS1 domain-containing protein

MacromoleculeName: BBS1 domain-containing protein / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Bos taurus (cattle) / Organ: Eye / Tissue: Retina
Molecular weightTheoretical: 64.939141 KDa
SequenceString: MAATSSSDSD GGKGESEANS KWLDSLSDSM ANIHTFSACL ALADFHGDGE YKLAMGDLGP DGRQPRLKVL KGHTLVSQKP LPDLPAAAV TFLMASHEPR TPALAIASGP CVYVYKNLKP YFKFSLPSLP TNPLEQDLWN QAKEDQIDPL TLKEMLEGIR E KAEVPLSV ...String:
MAATSSSDSD GGKGESEANS KWLDSLSDSM ANIHTFSACL ALADFHGDGE YKLAMGDLGP DGRQPRLKVL KGHTLVSQKP LPDLPAAAV TFLMASHEPR TPALAIASGP CVYVYKNLKP YFKFSLPSLP TNPLEQDLWN QAKEDQIDPL TLKEMLEGIR E KAEVPLSV QSLRFLPLEL SEMEAFVNQH KSKSIRRQTV ITTMTTLKKN LADEDAVSCL VLGTENKELL VLDPEAFTIL AK MSLPSVP AFLEASGQFD VEFRLAAACR NGSIYILRRD SKRPKYCIEL GAQPVGLVGV HKVLVVGSNQ DSLHGFTYKG KRL WTVQMP AAILAMNLLE QHSRGLQAVM AALANEEVRI YHDKVLLNVI RTPEAVTSLC FGRYGREDNT LIMTTLGGGL IIKI LKRTA VFAEGGGEAG PPPSQAIKLN VPRKTRLYVD QTLREREAGT AMHRTFQADL YLLRLRAARA YVQALESSLS PVSLT AREP LKLHAVVQGL GPTFKLTLHL QNTSTARPIL GLVVCFLYNE VLYALPRAFF KVPLLVPGLN YPLETFVKSL SDKGIS DII KVLVLREGQS TPLLSAHINM PMSEGLAAD

UniProtKB: Bardet-Biedl syndrome 1

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Macromolecule #3: Bardet-Biedl syndrome 2 protein homolog

MacromoleculeName: Bardet-Biedl syndrome 2 protein homolog / type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Bos taurus (cattle) / Organ: Eye / Tissue: Retina
Molecular weightTheoretical: 79.911484 KDa
SequenceString: MLQPVFTLKL RHKISPRMVA VGRYDGTHPC LAAATQAGKV FIHNPHSRSQ HLGAPRVLQS PLESDVSLLN INQTVSCLTA GVLNPELGY DALLVGTQTN LLAYDVYNNS DLFYREVADG ASAIVLGTLG DITSPLAIIG GNCALQGFNH EGNDLFWTVT G DNVHSLAL ...String:
MLQPVFTLKL RHKISPRMVA VGRYDGTHPC LAAATQAGKV FIHNPHSRSQ HLGAPRVLQS PLESDVSLLN INQTVSCLTA GVLNPELGY DALLVGTQTN LLAYDVYNNS DLFYREVADG ASAIVLGTLG DITSPLAIIG GNCALQGFNH EGNDLFWTVT G DNVHSLAL CDFDGDGKKE LLVGSEDFDI RVFKEDEIVA EMSETEIITS LCPMYGSRFG YALSNGTVGV YDKTARYWRI KS KNQAMSI HAFDLNSDGV CELITGWSNG KVDARSDRTG EVIFKDNFSS AIAGVVEGDY RMEGCQQLIC CSVDGEIRGY LPG TAEMRG NLMDISVEQD LIRELSQKKQ NLLLELRNYE ENAKAELSSP LNEADGHRGV IPANTKHHTA LSVSLGSEAQ AAHA ELCIS TSNDTIIRAV LIFAEGVFAG ESHVVHPSVH HLSSSVRIPI TPPKDIPVDL HLKTFVGYRS STQFHVFELT RQLPR FSMY ALTSPDPASE PLSYVNFIIA ERAQRVVMWL NQNFLLPEDT NIQNAPFQVC FTSLRNGGQL YIKIKLSGEI TVNTDD IDL AGDIIQSMAS FFAIEDLQVE ADFPVYFEEL RKVLVKVDEY HSVHQKLSAD MADNSNLIRS LLVQAEDARL MRDMKTM KN RYKELYDLNK DLLNGYKIRC NNHTELLGSL KAVNQAIQRA GHLRVGKPKN QVITACRDAI RSNNINMLFR IMRVGTAS S

UniProtKB: Bardet-Biedl syndrome 2 protein homolog

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Macromolecule #4: Bardet-Biedl syndrome 4 protein homolog

MacromoleculeName: Bardet-Biedl syndrome 4 protein homolog / type: protein_or_peptide / ID: 4 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Bos taurus (cattle) / Organ: Eye / Tissue: Retina
Molecular weightTheoretical: 58.289133 KDa
SequenceString: MAEEKLSART QLPVSAESQK PVLKKAPEFP ILEKQNWLIH LYYIQKDYEA CKAVIKEQLQ ETHGLCEYAI YVQALIFRLE GNIQESLRL FQMCAFLSPQ CADNLKQVAR SLFLLGKHKA AIEVYNEAAK LNQKDWEICH NLGVCYIYLK QFDKAQDQLH N ALHLNRHD ...String:
MAEEKLSART QLPVSAESQK PVLKKAPEFP ILEKQNWLIH LYYIQKDYEA CKAVIKEQLQ ETHGLCEYAI YVQALIFRLE GNIQESLRL FQMCAFLSPQ CADNLKQVAR SLFLLGKHKA AIEVYNEAAK LNQKDWEICH NLGVCYIYLK QFDKAQDQLH N ALHLNRHD LTYIMLGKIF LLKGDLDKAI EIYKKAVEFS PENTELLTTL GLLYLQLGIY QKAFEHLGNT LTYDPTNYKA IL AAGSMMQ THGDFDVALT KYKVVACAVI ESPPLWNNIG MCFFGKKKYV AAISCLKRAN YLAPLDWKIL YNLGLVHLTM QQY ASAFHF LSAAINFQPK MGELYMLLAV ALTNLEDSEN AKRAYEEAVR LDKCNPLVNL NYAVLLYNQG EKRDALAQYQ EMEK KVNLL KYSSSLEFDP EMVEVAQKLG AALQVGEALV WTKPVKDPKS KHQTASTSKA AGFQQPLGSN QALGQAMSSA ATCRK LSSG AGGTSQLTKP PSLPLEPEPT VEAQPTEASA QTREK

UniProtKB: Bardet-Biedl syndrome 4 protein homolog

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Macromolecule #5: Bardet-Biedl syndrome 5 protein homolog

MacromoleculeName: Bardet-Biedl syndrome 5 protein homolog / type: protein_or_peptide / ID: 5 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Bos taurus (cattle) / Organ: Eye / Tissue: Retina
Molecular weightTheoretical: 38.880984 KDa
SequenceString: MSVLDALWED RDVRFDVSSQ QMKTRPGEVL IDCLDSVEDT KGNNGDRGRL LVTNLRIVWH SLALPRVNLS IGYNCILNIT TRTANSKLR GQTEALYVLT KCNSTRFEFI FTNLVPGSPR LYTSLIAVHR AYETSKMYRD FKLRSALIQN KQLRLLPQEN V YNKINGVW ...String:
MSVLDALWED RDVRFDVSSQ QMKTRPGEVL IDCLDSVEDT KGNNGDRGRL LVTNLRIVWH SLALPRVNLS IGYNCILNIT TRTANSKLR GQTEALYVLT KCNSTRFEFI FTNLVPGSPR LYTSLIAVHR AYETSKMYRD FKLRSALIQN KQLRLLPQEN V YNKINGVW NLSSDQGNLG TFFITNVRIV WHANMNDSFN VSIPYLQIRS VKIRDSKFGL ALVIESSQQS GGYVLGFKID PV EKLQESV KEINSLHKVY SANPIFGVDY EMEEKPQPLE ALTVKQIQDD VEIDSDDHTD AFVAYFADGN KQQDREPVFS EEL GLAIEK LKDGFTLQGL WEVMN

UniProtKB: Bardet-Biedl syndrome 5 protein homolog

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Macromolecule #6: Bardet-Biedl syndrome 7 protein homolog

MacromoleculeName: Bardet-Biedl syndrome 7 protein homolog / type: protein_or_peptide / ID: 6 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Bos taurus (cattle) / Organ: Eye / Tissue: Retina
Molecular weightTheoretical: 80.471375 KDa
SequenceString: MDLNLNRADY LQVGVTSQKT MKLLPASKHR ATQKVVVGDH DGIVMCFGMK KGEAVTVFKT LPGQKIARLE LGGALNTPQE KIFIAAGSE IRGFTKRGKQ FLSFETNLTE SIKAMHISGS DLFLSASYIY NHYCDCKDQH YYLSGDKIND VICLPVERLL R EVPVLACQ ...String:
MDLNLNRADY LQVGVTSQKT MKLLPASKHR ATQKVVVGDH DGIVMCFGMK KGEAVTVFKT LPGQKIARLE LGGALNTPQE KIFIAAGSE IRGFTKRGKQ FLSFETNLTE SIKAMHISGS DLFLSASYIY NHYCDCKDQH YYLSGDKIND VICLPVERLL R EVPVLACQ DRVLRVLQGS DVTYEIEVPG PPTVLALHNG NGGDSGEDLL FGTSDGKLGL IQITTSKPIH KWEIRNEKKR GG ILCVDSF DIVGDGVKDL LVGRDDGMVE VYGFDNANEP VLRFDHTLSE SVTSIQGGCV GKDGYDEIVV STYSGWITGL TTE PVHKES GPGEELKFNQ EMQNKISSLR SELEQLQYKV LQEREKYQQS SQSSKAKSAV PSFSVNDKFT LNKDDASYSL ILEV QTAID NVLIQSDVPI DLLDVDKNSA VVSFSSCDSE SNDNFLLATY RCQANTTRLE LKIRSIEGQY GTLQAYVTPR IQPKT CQVR QYHIKPLSLH QRTHFIDHDR PMNTLTLTGQ FSFSELHSWV VFCMPEVPEK PPAGECVTFY FQNTFLDTQL ESTYRK GEG VFKSDNISTI SILKDVLSKE ATKRKINLNI SYEINEVSVK HTLKLIHPKL EYQLLLAKKV QLIDALKELQ VHEGNTN FL IPEYRCILEE ADHLQEEYKK QPAHLERLYG MITDLFIDKF KFKGTNVKTK VPLLLEILDS YDQNALIAFF DAA

UniProtKB: Bardet-Biedl syndrome 7 protein homolog

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Macromolecule #7: Tetratricopeptide repeat domain 8

MacromoleculeName: Tetratricopeptide repeat domain 8 / type: protein_or_peptide / ID: 7 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Bos taurus (cattle) / Organ: Eye / Tissue: Retina
Molecular weightTheoretical: 56.686406 KDa
SequenceString: MEPLLLAWSY FRRRRFQLCA DLCTQMLEKS PCDQAAWILK ARALTEMVYV DEIDVDEEGI AEMILDENAI AQVPRPGTSL KLPGTNQTG GPSPAVRPVT QAGRPITGFL RPSTQSGRPG TIEQAIKTPR TAYTARPIAS SSGRFVRLGT ASMLTSPDGP F INLSRLNL ...String:
MEPLLLAWSY FRRRRFQLCA DLCTQMLEKS PCDQAAWILK ARALTEMVYV DEIDVDEEGI AEMILDENAI AQVPRPGTSL KLPGTNQTG GPSPAVRPVT QAGRPITGFL RPSTQSGRPG TIEQAIKTPR TAYTARPIAS SSGRFVRLGT ASMLTSPDGP F INLSRLNL AKYAQKPKLA KALFEYIFHH ENDVKTALDL AALSTEHSQY KDWWWKVQIG KCYYRLGLYR EAEKQFKSAL KQ QEMVDTF LYLAKVYISL DQPLTALNLF KQGLDKFPGE VTLLCGIARI YEEMNNISSA TEYYKEVLKQ DNTHVEAIAC IGS NHFYTD QPEVALRFYR RLLQMGVYNC QLFNNLGLCC FYAQQYDMTL TSFERALSLA ENEEEVADVW YNLGHVAVGT GDTN LAHQC FRLALVSNNQ HAEAYNNLAV LEMRRGHVEQ AKALLQTASS LAPHMYEPHF NFATISDKIG DLQRSYAAAK KSEAA FPDH VDTQHLIKQL EQHFAML

UniProtKB: Tetratricopeptide repeat domain 8

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Macromolecule #8: Bardet-Biedl syndrome 9

MacromoleculeName: Bardet-Biedl syndrome 9 / type: protein_or_peptide / ID: 8 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Bos taurus (cattle) / Organ: Eye / Tissue: Retina
Molecular weightTheoretical: 99.230914 KDa
SequenceString: MSLFKARDWW STVLGDKEEF DQGCLCLADV DNTGNGQDKI IVGSFMGYLR IFNPHPVKTG DGAQAEDLLL EVHLRDPILQ VEVGKFVSG TEMLHLAVLH SRKLCVYSVS GTLGNVEHGN QYQIKLMYEH NLQRTACNMT YGSFGGVKGR DLICIQSVDG M LMVFEQES ...String:
MSLFKARDWW STVLGDKEEF DQGCLCLADV DNTGNGQDKI IVGSFMGYLR IFNPHPVKTG DGAQAEDLLL EVHLRDPILQ VEVGKFVSG TEMLHLAVLH SRKLCVYSVS GTLGNVEHGN QYQIKLMYEH NLQRTACNMT YGSFGGVKGR DLICIQSVDG M LMVFEQES YAFGRFLPGS LLPGPLAYSS RTDSFITVSS CHQVESYKYQ VLAFATDADK RQETEQQKHG SGKRLVVDWT LN IGEQAID ICIVSFIQSA SSVFVLGERN FFCLKDNGQI QFMKKLDYSP SCFLPYCSVS EGTINTLIGN HNNMLHIYQD VTL KWATQL PHVPVAVRVG CLHDLKGVIV TLSDDGHLQC SYLGTDPSLF QAPKVESREL NYDELDMELK ELQKVIKNVN KSQD VWPLT EREDDLKVSA MVSPNFDSVS QATDVEVGAD LVPSVTVKVT LKNRVALQKI KLSIYVQPPL VLTGDQFTFE FMAPE MTRT VGFSVYLKGS YSPPELEGNA VVSYSRPTER NPDGIPRVSQ CKFRLPLKLV CLPGQPSKTA SHKLTIDTNK SPVSLL SLF PGFAKQSEDD QVNVMGFRFL GGSQVTLLAS KTSQRYRIQS EQFEDLWLIT NELIIRLQEY FEKQGIKDFT CSFSGSV PL EEYFELIDHH FELRINGEKL EELLSERAVQ FRAIQRRLLT RFKDKTPAPL QHLDTLLDGT YKQVIALADA VEENQDNL F QSFTRLKSAT HLVILLIGLW QKLSADQIAI LEAAFLPLQQ DTQELGWEET VDAALSHLLK TCLSKSSKEQ ALNLNSQLG IPKDTSQLKK HITLFCDRLA KGGRLCLSTD AAAPQTMVMP GGCATIPESD LEGRSIDQDS SELFTNHKHL MVETPVPEVS PLQGVTE

UniProtKB: Bardet-Biedl syndrome 9

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Macromolecule #9: ADP-ribosylation factor-like protein 6

MacromoleculeName: ADP-ribosylation factor-like protein 6 / type: protein_or_peptide / ID: 9 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Bos taurus (cattle)
Molecular weightTheoretical: 21.086486 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString:
MGLLDRLSGL LGLKKKEVHV LCLGLDNSGK TTIINKLKPS NAQSQDIVPT IGFSIQKFKS SSLSFTVFDM SGQGRYRNLW EHYYKEGQA IIFVIDSSDK LRMVVAKEEL RTLLNHPDIK HRRIPILFFA NKMDLRDALT SVKVSQLLCL EDIKDKPWHI C ASDAIKGE GLQEGVDWLQ DQIQSVKT

UniProtKB: ADP-ribosylation factor-like protein 6

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Macromolecule #10: CALCIUM ION

MacromoleculeName: CALCIUM ION / type: ligand / ID: 10 / Number of copies: 2 / Formula: CA
Molecular weightTheoretical: 40.078 Da

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Macromolecule #11: GUANOSINE-5'-TRIPHOSPHATE

MacromoleculeName: GUANOSINE-5'-TRIPHOSPHATE / type: ligand / ID: 11 / Number of copies: 1 / Formula: GTP
Molecular weightTheoretical: 523.18 Da
Chemical component information

ChemComp-GTP:
GUANOSINE-5'-TRIPHOSPHATE / GTP, energy-carrying molecule*YM / Guanosine triphosphate

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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

Concentration0.7 mg/mL
BufferpH: 7.5
Component:
ConcentrationNameFormula
20.0 mMHepes
220.0 mMNaClSodium chloride
5.0 mMMgCl2
4.0 mMbeta-mercaptoethanol2-Mercaptoethanol
1.0 mMGTP
GridSupport film - Material: CARBON / Support film - topology: HOLEY / Details: unspecified
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 293 K / Instrument: FEI VITROBOT MARK II / Details: Grids were blotted for 2 s with a -2 offset..

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 50.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 2.4 µm / Nominal defocus min: 1.1 µm / Nominal magnification: 81000
Specialist opticsEnergy filter - Name: GIF Bioquantum / Energy filter - Slit width: 25 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 grids imaged: 2 / Number real images: 9408 / Average exposure time: 4.0 sec. / Average electron dose: 56.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: OTHER
Details: An initial map for the BBSome was generated using RELION's implementation of the stochastic gradient descent algorithm using default parameters and a mask diameter of 280 Angstrom.
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.0)
Final 3D classificationNumber classes: 8 / Software - Name: RELION (ver. 3.0)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.0)
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Algorithm: BACK PROJECTION / Resolution.type: BY AUTHOR / Resolution: 3.5 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.0) / Number images used: 75201
FSC plot (resolution estimation)

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Atomic model buiding 1

DetailsDuring refinement, the resolution limit was set to 3.5 Angstrom. Secondary structure, Ramachandran and rotamer restraints were applied during refinement.
RefinementSpace: REAL / Protocol: OTHER / Overall B value: 43.6 / Target criteria: Correlation coefficient
Output model

PDB-6vbv:
Structure of the bovine BBSome:ARL6:GTP complex

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