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- PDB-7u06: Structure of the yeast TRAPPII-Rab11/Ypt32 complex in the closed/... -

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Basic information

Entry
Database: PDB / ID: 7u06
TitleStructure of the yeast TRAPPII-Rab11/Ypt32 complex in the closed/open state (composite structure)
Components
  • (Trafficking protein particle complex II-specific subunit ...) x 5
  • (Trafficking protein particle complex subunit ...) x 6
  • GTP-binding protein YPT32/YPT11
  • TRAPP-associated protein TCA17
KeywordsPROTEIN TRANSPORT / complex / GTPase / Guanosine Exchange Factor / GEF
Function / homology
Function and homology information


Anchoring of the basal body to the plasma membrane / beta-glucan biosynthetic process / RAB geranylgeranylation / TRAPPI protein complex / RAB GEFs exchange GTP for GDP on RABs / TRAPPII protein complex / TRAPPIII protein complex / TRAPP complex / early endosome to Golgi transport / COPII-mediated vesicle transport ...Anchoring of the basal body to the plasma membrane / beta-glucan biosynthetic process / RAB geranylgeranylation / TRAPPI protein complex / RAB GEFs exchange GTP for GDP on RABs / TRAPPII protein complex / TRAPPIII protein complex / TRAPP complex / early endosome to Golgi transport / COPII-mediated vesicle transport / cis-Golgi network membrane / cytoplasm to vacuole targeting by the Cvt pathway / cellular bud neck / protein localization to phagophore assembly site / intra-Golgi vesicle-mediated transport / cis-Golgi network / protein-containing complex localization / retrograde transport, endosome to Golgi / phagophore assembly site / cellular response to nitrogen starvation / exocytosis / positive regulation of macroautophagy / chromosome organization / endoplasmic reticulum to Golgi vesicle-mediated transport / vesicle-mediated transport / Neutrophil degranulation / macroautophagy / trans-Golgi network / cell wall organization / recycling endosome / autophagy / protein transport / protein-containing complex assembly / mitochondrial outer membrane / early endosome / endosome / Golgi membrane / GTPase activity / GTP binding / Golgi apparatus / endoplasmic reticulum / nucleus / cytosol / cytoplasm
Similarity search - Function
TRAPP II complex, Trs120 / TRAPP II complex TRAPPC10, C-terminal / Trafficking protein particle complex II-specific subunit 65 / Trafficking protein particle complex subunit TRAPPC10/Trs130 / Transport protein Trs120 or TRAPPC9, TRAPP II complex subunit / Trafficking protein particle complex subunit 10, TRAPPC10 / TRAPP trafficking subunit Trs65 / Trafficking protein particle complex subunit 2 / Sedlin, N-terminal conserved region / Trafficking protein particle complex subunit ...TRAPP II complex, Trs120 / TRAPP II complex TRAPPC10, C-terminal / Trafficking protein particle complex II-specific subunit 65 / Trafficking protein particle complex subunit TRAPPC10/Trs130 / Transport protein Trs120 or TRAPPC9, TRAPP II complex subunit / Trafficking protein particle complex subunit 10, TRAPPC10 / TRAPP trafficking subunit Trs65 / Trafficking protein particle complex subunit 2 / Sedlin, N-terminal conserved region / Trafficking protein particle complex subunit / Sybindin-like family / Sybindin-like family / TRAPP I complex, subunit 5 / TRAPP complex, Trs33 subunit / Bet3 family / Transport protein particle (TRAPP) component / Transport protein particle (TRAPP) component / : / NO signalling/Golgi transport ligand-binding domain superfamily / Longin-like domain superfamily / small GTPase Rab1 family profile. / Ran (Ras-related nuclear proteins) /TC4 subfamily of small GTPases / Rho (Ras homology) subfamily of Ras-like small GTPases / Ras subfamily of RAS small GTPases / Small GTPase / Ras family / Rab subfamily of small GTPases / Small GTP-binding protein domain / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
PALMITIC ACID / TRAPP-associated protein TCA17 / Trafficking protein particle complex II-specific subunit 65 / Trafficking protein particle complex subunit BET3 / Trafficking protein particle complex subunit 20 / GTP-binding protein YPT32/YPT11 / Trafficking protein particle complex subunit 31 / Trafficking protein particle complex subunit BET5 / Trafficking protein particle complex II-specific subunit 130 / Trafficking protein particle complex subunit 23 ...PALMITIC ACID / TRAPP-associated protein TCA17 / Trafficking protein particle complex II-specific subunit 65 / Trafficking protein particle complex subunit BET3 / Trafficking protein particle complex subunit 20 / GTP-binding protein YPT32/YPT11 / Trafficking protein particle complex subunit 31 / Trafficking protein particle complex subunit BET5 / Trafficking protein particle complex II-specific subunit 130 / Trafficking protein particle complex subunit 23 / Trafficking protein particle complex II-specific subunit 120 / Trafficking protein particle complex subunit 33
Similarity search - Component
Biological speciesSaccharomyces cerevisiae (brewer's yeast)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 4.2 Å
AuthorsBagde, S.R. / Fromme, J.C.
Funding support United States, 2items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R35GM136258 United States
National Science Foundation (NSF, United States)DMR-1719875 United States
CitationJournal: Sci Adv / Year: 2022
Title: Structure of a TRAPPII-Rab11 activation intermediate reveals GTPase substrate selection mechanisms.
Authors: Saket R Bagde / J Christopher Fromme /
Abstract: Rab1 and Rab11 are essential regulators of the eukaryotic secretory and endocytic recycling pathways. The transport protein particle (TRAPP) complexes activate these guanosine triphosphatases via ...Rab1 and Rab11 are essential regulators of the eukaryotic secretory and endocytic recycling pathways. The transport protein particle (TRAPP) complexes activate these guanosine triphosphatases via nucleotide exchange using a shared set of core subunits. The basal specificity of the TRAPP core is toward Rab1, yet the TRAPPII complex is specific for Rab11. A steric gating mechanism has been proposed to explain TRAPPII counterselection against Rab1. Here, we present cryo-electron microscopy structures of the 22-subunit TRAPPII complex from budding yeast, including a TRAPPII-Rab11 nucleotide exchange intermediate. The Trs130 subunit provides a "leg" that positions the active site distal to the membrane surface, and this leg is required for steric gating. The related TRAPPIII complex is unable to activate Rab11 because of a repulsive interaction, which TRAPPII surmounts using the Trs120 subunit as a "lid" to enclose the active site. TRAPPII also adopts an open conformation enabling Rab11 to access and exit from the active site chamber.
History
DepositionFeb 17, 2022Deposition site: RCSB / Processing site: RCSB
Revision 1.0Apr 27, 2022Provider: repository / Type: Initial release
Revision 1.1May 25, 2022Group: Database references / Category: citation
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.journal_volume / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.year

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
C: Trafficking protein particle complex II-specific subunit 65
D: TRAPP-associated protein TCA17
E: Trafficking protein particle complex subunit 33
F: Trafficking protein particle complex subunit BET3
G: Trafficking protein particle complex subunit BET5
H: Trafficking protein particle complex subunit 23
I: Trafficking protein particle complex subunit BET3
J: Trafficking protein particle complex subunit 31
K: Trafficking protein particle complex subunit 20
l: GTP-binding protein YPT32/YPT11
B: Trafficking protein particle complex II-specific subunit 130
A: Trafficking protein particle complex II-specific subunit 120
c: Trafficking protein particle complex II-specific subunit 65
d: TRAPP-associated protein TCA17
e: Trafficking protein particle complex subunit 33
f: Trafficking protein particle complex subunit BET3
g: Trafficking protein particle complex subunit BET5
h: Trafficking protein particle complex subunit 23
i: Trafficking protein particle complex subunit BET3
j: Trafficking protein particle complex subunit 31
k: Trafficking protein particle complex subunit 20
b: Trafficking protein particle complex II-specific subunit 130
a: Trafficking protein particle complex II-specific subunit 120
M: Trafficking protein particle complex II-specific subunit 130
m: Trafficking protein particle complex II-specific subunit 130
n: Trafficking protein particle complex II-specific subunit 65
N: Trafficking protein particle complex II-specific subunit 65
hetero molecules


Theoretical massNumber of molelcules
Total (without water)1,165,95031
Polymers1,164,92427
Non-polymers1,0264
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: gel filtration
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

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Trafficking protein particle complex II-specific subunit ... , 5 types, 10 molecules CcBbAaMmnN

#1: Protein Trafficking protein particle complex II-specific subunit 65 / TRAPP II-specific subunit 65 / Beta-glucan synthesis-associated protein TRS65 / Killer toxin- ...TRAPP II-specific subunit 65 / Beta-glucan synthesis-associated protein TRS65 / Killer toxin-resistance protein 11 / Transport protein particle 65 kDa subunit


Mass: 63434.473 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / References: UniProt: P32893
#10: Protein Trafficking protein particle complex II-specific subunit 130 / TRAPP II-specific subunit 130 / Transport protein particle 130 kDa subunit


Mass: 128424.250 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / References: UniProt: Q03660
#11: Protein Trafficking protein particle complex II-specific subunit 120 / TRAPP II-specific subunit 120 / Transport protein particle 120 kDa subunit


Mass: 147823.281 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / References: UniProt: Q04183
#12: Protein Trafficking protein particle complex II-specific subunit 130


Mass: 24953.623 Da / Num. of mol.: 2 / Fragment: N-terminal region / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast)
#13: Protein Trafficking protein particle complex II-specific subunit 65


Mass: 17889.996 Da / Num. of mol.: 2 / Fragment: N-terminal region / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast)

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Protein , 2 types, 3 molecules Ddl

#2: Protein TRAPP-associated protein TCA17 / 17 kDa TRAPP complex-associated protein


Mass: 17371.008 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / References: UniProt: P32613
#9: Protein GTP-binding protein YPT32/YPT11 / Rab GTPase YPT32


Mass: 25309.145 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharomyces cerevisiae (brewer's yeast)
Gene: YPT32, YPT11, YGL210W / Production host: Escherichia coli BL21 (bacteria) / Strain (production host): Rosetta 2 / References: UniProt: P51996

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Trafficking protein particle complex subunit ... , 6 types, 14 molecules EeFIfiGgHhJjKk

#3: Protein Trafficking protein particle complex subunit 33 / TRAPP subunit 33 / Transport protein particle 33 kDa subunit


Mass: 30786.176 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / References: UniProt: Q99394
#4: Protein
Trafficking protein particle complex subunit BET3 / TRAPP subunit BET3 / Transport protein particle 22 kDa subunit


Mass: 22152.445 Da / Num. of mol.: 4 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / References: UniProt: P36149
#5: Protein Trafficking protein particle complex subunit BET5 / TRAPP subunit BET5 / Transport protein particle 18 kDa subunit


Mass: 18453.875 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / References: UniProt: Q03630
#6: Protein Trafficking protein particle complex subunit 23 / TRAPP subunit 23 / Transport protein particle 23 kDa subunit


Mass: 24889.262 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / References: UniProt: Q03784
#7: Protein Trafficking protein particle complex subunit 31 / TRAPP subunit 31 / Transport protein particle 31 kDa subunit


Mass: 31755.689 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / References: UniProt: Q03337
#8: Protein Trafficking protein particle complex subunit 20 / TRAPP subunit 20 / Transport protein particle 20 kDa subunit


Mass: 19721.154 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Source: (natural) Saccharomyces cerevisiae (brewer's yeast) / References: UniProt: P38334

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Non-polymers , 1 types, 4 molecules

#14: Chemical
ChemComp-PLM / PALMITIC ACID


Mass: 256.424 Da / Num. of mol.: 4 / Source method: obtained synthetically / Formula: C16H32O2 / Feature type: SUBJECT OF INVESTIGATION

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Details

Has ligand of interestY

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

ComponentName: TRAPPII complex bound to Rab11/Ypt32 / Type: COMPLEX / Entity ID: #1-#13 / Source: MULTIPLE SOURCES
Molecular weightValue: 1.1 MDa / Experimental value: NO
Source (natural)Organism: Saccharomyces cerevisiae (brewer's yeast)
Source (recombinant)Organism: Escherichia coli BL21(DE3) (bacteria) / Strain: BL21(DE3)
Buffer solutionpH: 8
Buffer component
IDConc.NameBuffer-ID
110 mMTris1
2150 mMSodium chloride1
30.1 %CHAPS1
41 mMMagnesium acetate1
51 mMDTT1
SpecimenConc.: 5.6 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: GOLD / Grid mesh size: 300 divisions/in. / Grid type: UltrAuFoil R1.2/1.3
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277 K
Details: The sample was incubated on the grid for 10 seconds followed by blotting for 5 seconds before plunging in liquid ethane.

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Electron microscopy imaging

Experimental equipment
Model: Talos Arctica / Image courtesy: FEI Company
MicroscopyModel: FEI TALOS ARCTICA
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 200 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal magnification: 63000 X / Nominal defocus max: 2500 nm / Nominal defocus min: 800 nm / Cs: 2.7 mm / C2 aperture diameter: 50 µm / Alignment procedure: COMA FREE
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingAverage exposure time: 3.5 sec. / Electron dose: 53 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Num. of grids imaged: 3 / Num. of real images: 4998 / Details: Images were collected as 50 frame movies.
EM imaging opticsEnergyfilter name: GIF Bioquantum / Energyfilter slit width: 20 eV
Image scansWidth: 5760 / Height: 4092

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Processing

Software
NameVersionClassification
phenix.real_space_refine1.19.2_4158refinement
PHENIX1.19.2_4158refinement
EM software
IDNameVersionCategoryDetails
2EPUimage acquisition
4cryoSPARC3.2.0CTF correctionPatch CTF estimation (multi)
5RELION3.1CTF correction
8Cootmodel fitting
10PHENIXmodel refinement
11cryoSPARC3.2.0initial Euler assignment
12RELION3.1final Euler assignment
14PHENIX3D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 979187
SymmetryPoint symmetry: C1 (asymmetric)
3D reconstructionResolution: 4.2 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 149906
Details: The composite map was generated by combining the consensus dimer map (EMD-26233) with focused refinement maps deposited in the entries EMD-26234, EMD-26235, EMD-26236, EMD-26237, EMD-26238, ...Details: The composite map was generated by combining the consensus dimer map (EMD-26233) with focused refinement maps deposited in the entries EMD-26234, EMD-26235, EMD-26236, EMD-26237, EMD-26238, EMD-26239, EMD-26240, EMD-26241, EMD-26242, EMD-26243, EMD-26244, EMD-26245, EMD-26246, EMD-26247, EMD-26248, EMD-26249, EMD-26250, EMD-26251, EMD-26252 and EMD-26253 using Combine Focused Maps in Phenix.
Symmetry type: POINT
Atomic model buildingProtocol: OTHER / Space: REAL
Details: Model was rebuilt in coot and refined using phenix.real_space_refine.
Atomic model building
IDPDB-ID 3D fitting-ID
13CUE

3cue

1
23RWO

3rwo

1
33PR6

3pr6

1
RefinementCross valid method: NONE
Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2
Displacement parametersBiso mean: 104.06 Å2
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.004665054
ELECTRON MICROSCOPYf_angle_d0.817388071
ELECTRON MICROSCOPYf_chiral_restr0.047410188
ELECTRON MICROSCOPYf_plane_restr0.005611161
ELECTRON MICROSCOPYf_dihedral_angle_d5.96998721

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