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- EMDB-26716: Gea2-Arf1 activation intermediate complex (composite structure) -

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Basic information

Entry
Database: EMDB / ID: EMD-26716
TitleGea2-Arf1 activation intermediate complex (composite structure)
Map dataGea2-Arf1 activation intermediate complex (composite structure)
Sample
  • Complex: Gea2 bound to Arf1
    • Complex: GEA2 isoform 1
      • Protein or peptide: GEA2 isoform 1
    • Complex: ARF1 isoform 1
      • Protein or peptide: ARF1 isoform 1
KeywordsGEF / PROTEIN TRANSPORT
Function / homology
Function and homology information


cellular localization / regulation of ARF protein signal transduction / vesicle-mediated transport / guanyl-nucleotide exchange factor activity / protein transport / GTPase activity / GTP binding / Golgi apparatus
Similarity search - Function
Mon2/Sec7/BIG1-like, HUS domain / Mon2/Sec7/BIG1-like, HUS domain / Sec7 domain / Sec7, C-terminal domain superfamily / Sec7 domain superfamily / Sec7 domain / SEC7 domain profile. / Sec7 domain / ADP-ribosylation factor 1-5 / Small GTPase superfamily, ARF type ...Mon2/Sec7/BIG1-like, HUS domain / Mon2/Sec7/BIG1-like, HUS domain / Sec7 domain / Sec7, C-terminal domain superfamily / Sec7 domain superfamily / Sec7 domain / SEC7 domain profile. / Sec7 domain / ADP-ribosylation factor 1-5 / Small GTPase superfamily, ARF type / Sar1p-like members of the Ras-family of small GTPases / Small GTPase superfamily, ARF/SAR type / ADP-ribosylation factor family / Small GTP-binding protein domain / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
ADP-ribosylation factor / GEA2 isoform 1
Similarity search - Component
Biological speciesSaccharomyces cerevisiae (brewer's yeast)
Methodsingle particle reconstruction / cryo EM / Resolution: 4.2 Å
AuthorsMuccini A / Fromme JC
Funding support United States, 1 items
OrganizationGrant numberCountry
National Institutes of Health/National Cancer Institute (NIH/NCI)R35GM136258 United States
CitationJournal: Cell Rep / Year: 2022
Title: Structural basis for activation of Arf1 at the Golgi complex.
Authors: Arnold J Muccini / Margaret A Gustafson / J Christopher Fromme /
Abstract: The Golgi complex is the central sorting station of the eukaryotic secretory pathway. Traffic through the Golgi requires activation of Arf guanosine triphosphatases that orchestrate cargo sorting and ...The Golgi complex is the central sorting station of the eukaryotic secretory pathway. Traffic through the Golgi requires activation of Arf guanosine triphosphatases that orchestrate cargo sorting and vesicle formation by recruiting an array of effector proteins. Arf activation and Golgi membrane association is controlled by large guanine nucleotide exchange factors (GEFs) possessing multiple conserved regulatory domains. Here we present cryoelectron microscopy (cryoEM) structures of full-length Gea2, the yeast paralog of the human Arf-GEF GBF1, that reveal the organization of these regulatory domains and explain how Gea2 binds to the Golgi membrane surface. We find that the GEF domain adopts two different conformations compatible with different stages of the Arf activation reaction. The structure of a Gea2-Arf1 activation intermediate suggests that the movement of the GEF domain primes Arf1 for membrane insertion upon guanosine triphosphate binding. We propose that conformational switching of Gea2 during the nucleotide exchange reaction promotes membrane insertion of Arf1.
History
DepositionApr 22, 2022-
Header (metadata) releaseAug 24, 2022-
Map releaseAug 24, 2022-
UpdateMay 1, 2024-
Current statusMay 1, 2024Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_26716.png

Downloads & links

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Map

FileDownload / File: emd_26716.map.gz / Format: CCP4 / Size: 103 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationGea2-Arf1 activation intermediate complex (composite structure)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesX (Sec.)Y (Row.)Z (Col.)
1.65 Å/pix.
x 300 pix.
= 495.36 Å		1.65 Å/pix.
x 300 pix.
= 495.36 Å		1.65 Å/pix.
x 300 pix.
= 495.36 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.6512 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 5.6
Minimum - Maximum-134.720100000000002 - 142.729230000000001
Average (Standard dev.)0.003393386 (±1.5517825)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderZYX
Origin000
Dimensions300300300
Spacing300300300
CellA=B=C: 495.36002 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_26716_msk_1.map
Projections & Slices
AxesZYX

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Additional map: Gea2-Arf1 activation intermediate complex (composite structure)

Fileemd_26716_additional_1.map
AnnotationGea2-Arf1 activation intermediate complex (composite structure)
Projections & Slices
AxesZYX

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Additional map: Gea2-Arf1 activation intermediate complex (composite structure)

Fileemd_26716_additional_2.map
AnnotationGea2-Arf1 activation intermediate complex (composite structure)
Projections & Slices
AxesZYX

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Histogram

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Additional map: Gea2-Arf1 activation intermediate complex (composite structure)

Fileemd_26716_additional_3.map
AnnotationGea2-Arf1 activation intermediate complex (composite structure)
Projections & Slices
AxesZYX

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Additional map: Gea2-Arf1 activation intermediate complex (composite structure)

Fileemd_26716_additional_4.map
AnnotationGea2-Arf1 activation intermediate complex (composite structure)
Projections & Slices
AxesZYX

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Half map: Gea2-Arf1 activation intermediate complex (composite structure)

Fileemd_26716_half_map_1.map
AnnotationGea2-Arf1 activation intermediate complex (composite structure)
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Gea2-Arf1 activation intermediate complex (composite structure)

Fileemd_26716_half_map_2.map
AnnotationGea2-Arf1 activation intermediate complex (composite structure)
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Gea2 bound to Arf1

EntireName: Gea2 bound to Arf1
Components
  • Complex: Gea2 bound to Arf1
    • Complex: GEA2 isoform 1
      • Protein or peptide: GEA2 isoform 1
    • Complex: ARF1 isoform 1
      • Protein or peptide: ARF1 isoform 1

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Supramolecule #1: Gea2 bound to Arf1

SupramoleculeName: Gea2 bound to Arf1 / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all

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Supramolecule #2: GEA2 isoform 1

SupramoleculeName: GEA2 isoform 1 / type: complex / ID: 2 / Parent: 1 / Macromolecule list: #1
Source (natural)Organism: Saccharomyces cerevisiae (brewer's yeast)

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Supramolecule #3: ARF1 isoform 1

SupramoleculeName: ARF1 isoform 1 / type: complex / ID: 3 / Parent: 1 / Macromolecule list: #2
Source (natural)Organism: Saccharomyces cerevisiae (brewer's yeast)

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Macromolecule #1: GEA2 isoform 1

MacromoleculeName: GEA2 isoform 1 / type: protein_or_peptide / ID: 1 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Saccharomyces cerevisiae (brewer's yeast)
Molecular weightTheoretical: 165.846844 KDa
Recombinant expressionOrganism: Komagataella pastoris (fungus)
SequenceString: MSDREFVTVD PVTIIIKECI NLSTAMRKYS KFTSQSGVAA LLGGGSEIFS NQDDYLAHTF NNLNTNKHND PFLSGFIQLR LMLNKLKNL DNIDSLTILQ PFLLIVSTSS ISGYITSLAL DSLQKFFTLN IINESSQNYI GAHRATVNAL THCRFEGSQQ L SDDSVLLK ...String:
MSDREFVTVD PVTIIIKECI NLSTAMRKYS KFTSQSGVAA LLGGGSEIFS NQDDYLAHTF NNLNTNKHND PFLSGFIQLR LMLNKLKNL DNIDSLTILQ PFLLIVSTSS ISGYITSLAL DSLQKFFTLN IINESSQNYI GAHRATVNAL THCRFEGSQQ L SDDSVLLK VVFLLRSIVD SPYGDLLSNS IIYDVLQTIL SLACNNRRSE VLRNAAQSTM IAVTVKIFSK LKTIEPVNVN QI YINDESY TNDVLKADTI GTNVESKEEG SQEDPIGMKV NNEEAISEDD GIEEEHIHSE KSTNGAEQLD IVQKTTRSNS RIQ AYADDN YGLPVVRQYL NLLLSLIAPE NELKHSYSTR IFGLELIQTA LEISGDRLQL YPRLFTLISD PIFKSILFII QNTT KLSLL QATLQLFTTL VVILGNNLQL QIELTLTRIF SILLDDGTAN NSSSENKNKP SIIKELLIEQ ISILWTRSPS FFTST FINF DCNLDRADVS INFLKALTKL ALPESALTTT ESVPPICLEG LVSLVDDMFD HMKDIDREEF GRQKNEMEIL KKRDRK TEF IECTNAFNEK PKKGIPMLIE KGFIASDSDK DIAEFLFNNN NRMNKKTIGL LLCHPDKVSL LNEYIRLFDF SGLRVDE AI RILLTKFRLP GESQQIERII EAFSSAYCEN QDYDPSKISD NAEDDISTVQ PDADSVFILS YSIIMLNTDL HNPQVKEH M SFEDYSGNLK GCCNHKDFPF WYLDRIYCSI RDKEIVMPEE HHGNEKWFED AWNNLISSTT VITEIKKDTQ SVMDKLTPL ELLNFDRAIF KQVGPSIVST LFNIYVVASD DHISTRMITS LDKCSYISAF FDFKDLFNDI LNSIAKGTTL INSSHDDELS TLAFEYGPM PLVQIKFEDT NTEIPVSTDA VRFGRSFKGQ LNTVVFFRII RRNKDPKIFS KELWLNIVNI ILTLYEDLIL S PDIFPDLQ KRLKLSNLPK PSPEISINKS KESKGLLSTF ASYLKGDEEP TEEEIKSSKK AMECIKSSNI AASVFGNESN IT ADLIKTL LDSAKTEKNA DNSRYFEAEL LFIIELTIAL FLFCKEEKEL GKFILQKVFQ LSHTKGLTKR TVRRMLTYKI LLI SLCADQ TEYLSKLIND ELLKKGDIFT QKFFATNQGK EFLKRLFSLT ESEFYRGFLL GNENFWKFLR KVTAMKEQSE SIFE YLNES IKTDSNILTN ENFMWVLGLL DEISSMGAVG NHWEIEYKKL TESGHKIDKE NPYKKSIELS LKSIQLTSHL LEDNN DLRK NEIFAIIQAL AHQCINPCKQ ISEFAVVTLE QTLINKIEIP TNEMESVEEL IEGGLLPLLN SSETQEDQKI LISSIL TII SNVYLHYLKL GKTSNETFLK ILSIFNKFVE DSDIEKKLQQ LILDKKSIEK GNGSSSHGSA HEQTPESNDV EIEATAP ID DNTDDDNKPK LSDVEKD

UniProtKB: GEA2 isoform 1

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Macromolecule #2: ARF1 isoform 1

MacromoleculeName: ARF1 isoform 1 / type: protein_or_peptide / ID: 2 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Saccharomyces cerevisiae (brewer's yeast)
Molecular weightTheoretical: 20.552438 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString:
MGLFASKLFS NLFGNKEMRI LMVGLDGAGK TTVLYKLKLG EVITTIPTIG FNVETVQYKN ISFTVWDVGG QDRIRSLWRH YYRNTEGVI FVVDSNDRSR IGEAREVMQR MLNEDELRNA AWLVFANKQD LPEAMSAAEI TEKLGLHSIR NRPWFIQATC A TSGEGLYE GLEWLSNSLK NST

UniProtKB: ADP-ribosylation factor

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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
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TALOS ARCTICA
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average electron dose: 1.0 e/Å2
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.0 µm / Nominal defocus min: 1.0 µm
Experimental equipment
Model: Talos Arctica / Image courtesy: FEI Company

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Image processing

Startup modelType of model: INSILICO MODEL
Final reconstructionResolution.type: BY AUTHOR / Resolution: 4.2 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 391360
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD

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