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- EMDB-19410: COPII inner coat with Sed5 cargo bound -

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Basic information

Entry
Database: EMDB / ID: EMD-19410
TitleCOPII inner coat with Sed5 cargo bound
Map dataLocal resolution filtered (LocRes) consensus map.
Sample
  • Complex: Membrane-bound COPII inner coat with Sed5 bound
    • Protein or peptide: Sec23
    • Protein or peptide: Sec24
    • Protein or peptide: Sar1
    • Protein or peptide: Sed5
KeywordsCOPII / coat / PROTEIN TRANSPORT
Biological speciesSaccharomyces cerevisiae (brewer's yeast)
Methodsubtomogram averaging / cryo EM / Resolution: 4.13 Å
AuthorsZanetti G / Pyle E
Funding supportEuropean Union, United Kingdom, 4 items
OrganizationGrant numberCountry
European Research Council (ERC)852915European Union
Biotechnology and Biological Sciences Research Council (BBSRC)BB/T002670/1 United Kingdom
Wellcome Trust202679/Z/16/Z United Kingdom
Wellcome Trust206166/Z/17/Z United Kingdom
CitationJournal: Nat Struct Mol Biol / Year: 2025
Title: Cryo-electron tomography reveals how COPII assembles on cargo-containing membranes.
Authors: Euan Pyle / Elizabeth A Miller / Giulia Zanetti /
Abstract: Proteins traverse the eukaryotic secretory pathway through membrane trafficking between organelles. The coat protein complex II (COPII) mediates the anterograde transport of newly synthesized ...Proteins traverse the eukaryotic secretory pathway through membrane trafficking between organelles. The coat protein complex II (COPII) mediates the anterograde transport of newly synthesized proteins from the endoplasmic reticulum, engaging cargoes with a wide range of size and biophysical properties. The native architecture of the COPII coat and how cargo might influence COPII carrier morphology remain poorly understood. Here we reconstituted COPII-coated membrane carriers using purified Saccharomyces cerevisiae proteins and cell-derived microsomes as a native membrane source. Using cryo-electron tomography with subtomogram averaging, we demonstrate that the COPII coat binds cargo and forms largely spherical vesicles from native membranes. We reveal the architecture of the inner and outer coat layers and shed light on how spherical carriers are formed. Our results provide insights into the architecture and regulation of the COPII coat and advance our current understanding of how membrane curvature is generated.
History
DepositionJan 15, 2024-
Header (metadata) releaseJan 31, 2024-
Map releaseJan 31, 2024-
UpdateMar 26, 2025-
Current statusMar 26, 2025Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_19410.png

Downloads & links

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Map

FileDownload / File: emd_19410.map.gz / Format: CCP4 / Size: 28.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationLocal resolution filtered (LocRes) consensus map.
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.33 Å/pix.
x 196 pix.
= 260.484 Å		1.33 Å/pix.
x 196 pix.
= 260.484 Å		1.33 Å/pix.
x 196 pix.
= 260.484 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.329 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.178
Minimum - Maximum-0.7330536 - 1.0490905
Average (Standard dev.)-0.00000013216831 (±0.06372396)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions196196196
Spacing196196196
CellA=B=C: 260.484 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_19410_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half map 1.

Fileemd_19410_half_map_1.map
AnnotationHalf map 1.
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half map 2.

Fileemd_19410_half_map_2.map
AnnotationHalf map 2.
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Membrane-bound COPII inner coat with Sed5 bound

EntireName: Membrane-bound COPII inner coat with Sed5 bound
Components
  • Complex: Membrane-bound COPII inner coat with Sed5 bound
    • Protein or peptide: Sec23
    • Protein or peptide: Sec24
    • Protein or peptide: Sar1
    • Protein or peptide: Sed5

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Supramolecule #1: Membrane-bound COPII inner coat with Sed5 bound

SupramoleculeName: Membrane-bound COPII inner coat with Sed5 bound / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Details: COPII inner coat bound to GUV membrane with Sed5 bound.
Source (natural)Organism: Saccharomyces cerevisiae (brewer's yeast)
Molecular weightTheoretical: 247.532 KDa

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Macromolecule #1: Sec23

MacromoleculeName: Sec23 / type: protein_or_peptide / ID: 1 / Enantiomer: LEVO
Source (natural)Organism: Saccharomyces cerevisiae (brewer's yeast)
Recombinant expressionOrganism: Spodoptera frugiperda (fall armyworm)
SequenceString: DFETNEDING VRFTWNVFP S TRSDANSN VV PVGCLYT PLK EYDELN VAPY NPVVC SGPHC KSIL NPYCVI DPR NSSWSCP IC NSRNHLPP Q YTNLSQENM PLELQSTTIE YITNKPVTV P PIFFFVVD LT SETENLD SLK ESIITS LSLL PPNAL ...String:
DFETNEDING VRFTWNVFP S TRSDANSN VV PVGCLYT PLK EYDELN VAPY NPVVC SGPHC KSIL NPYCVI DPR NSSWSCP IC NSRNHLPP Q YTNLSQENM PLELQSTTIE YITNKPVTV P PIFFFVVD LT SETENLD SLK ESIITS LSLL PPNAL IGLIT YGNV VQLHDL SSE TIDRCNV FR GDREYQLE A LTEMLTGQK PTGPGGAASH LPNAMNKVT P FSLNRFFL PL EQVEFKL NQL LENLSP DQWS VPAGH RPLRA TGSA LNIASL LLQ GCYKNIP AR IILFASGP G TVAPGLIVN SELKDPLRSH HDIDSDHAQ H YKKACKFY NQ IAQRVAA NGH TVDIFA GCYD QIGMS EMKQL TDST GGVLLL TDA FSTAIFK QS YLRLFAKD E EGYLKMAFN GNMAVKTSKD LKVQGLIGH A SAVKKTDA NN ISESEIG IGA TSTWKM ASLS PYHSY AIFFE IANT AANSNP MMS APGSADR PH LAYTQFIT T YQHSSGTNR IRVTTVANQL LPFGTPAIA A SFDQEAAA VL MARIAVH KAE TDDGAD VIRW LDRTL IKLCQ KYAD YNKDDP QSF RLAPNFS LY PQFTYYLR R SQFLSVFNN SPDETAFYRH IFTREDTTN S LIMIQPTL TS FSMEDDP QPV LLDSIS VKPN TILLL DTFFF ILIY HGEQIA QWR KAGYQDD PQ YADFKALL E EPKLEAAEL LVDRFPLPRF IDTEAGGSQ A RFLLSKLN PS DNYQDMA RGG STIVLT DDVS LQNFM THLQQ VAVS GQA

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Macromolecule #2: Sec24

MacromoleculeName: Sec24 / type: protein_or_peptide / ID: 2 / Enantiomer: LEVO
Source (natural)Organism: Saccharomyces cerevisiae (brewer's yeast)
Recombinant expressionOrganism: Spodoptera frugiperda (fall armyworm)
SequenceString: MSHHKKRVYP QAQLQYGQN A TPLQQPAQ FM PPQDPAA AGM SYGQMG MPPQ GAVPS MGQQQ FLTP AQEQLH QQI DQATTSM ND MHLHNVPL V DPNAYMQPQ VPVQMGTPLQ QQQQPMAAP A YGQPSAAM GQ NMRPMNQ LYP IDLLTE LPPP ITDLT ...String:
MSHHKKRVYP QAQLQYGQN A TPLQQPAQ FM PPQDPAA AGM SYGQMG MPPQ GAVPS MGQQQ FLTP AQEQLH QQI DQATTSM ND MHLHNVPL V DPNAYMQPQ VPVQMGTPLQ QQQQPMAAP A YGQPSAAM GQ NMRPMNQ LYP IDLLTE LPPP ITDLT LPPPP LVIP PERMLV PSE LSNASPD YI RSTLNAVP K NSSLLKKSK LPFGLVIRPY QHLYDDIDP P PLNEDGLI VR CRRCRSY MNP FVTFIE QGRR WRCNF CRLAN DVPM QMDQSD PND PKSRYDR NE IKCAVMEY M APKEYTLRQ PPPATYCFLI DVSQSSIKS G LLATTINT LL QNLDSIP NHD ERTRIS ILCV DNAIH YFKIP LDSE NNEESA DQI NMMDIAD LE EPFLPRPN S MVVSLKACR QNIETLLTKI PQIFQSNLI T NFALGPAL KS AYHLIGG VGG KIIVVS GTLP NLGIG KLQRR NESG VVNTSK ETA QLLSCQD SF YKNFTIDC S KVQITVDLF LASEDYMDVA SLSNLSRFT A GQTHFYPG FS GKNPNDI VKF STEFAK HISM DFCME TVMRA RGST GLRMSR FYG HFFNRSS DL CAFSTMPR D QSYLFEVNV DESIMADYCY VQVAVLLSL N NSQRRIRI IT LAMPTTE SLA EVYASA DQLA IASFY NSKAV EKAL NSSLDD ARV LINKSVQ DI LATYKKEI V VSNTAGGAP LRLCANLRMF PLLMHSLTK H MAFRSGIV PS DHRASAL NNL ESLPLK YLIK NIYPD VYSLH DMAD EAGLPV QTE DGEATGT IV LPQPINAT S SLFERYGLY LIDNGNELFL WMGGDAVPA L VFDVFGTQ DI FDIPIGK QEI PVVENS EFNQ RVRNI INQLR NHDD VITYQS LYI VRGASLS EP VNHASARE V ATLRLWASS TLVEDKILNN ESYREFLQI M KARISK

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Macromolecule #3: Sar1

MacromoleculeName: Sar1 / type: protein_or_peptide / ID: 3 / Enantiomer: LEVO
Source (natural)Organism: Saccharomyces cerevisiae (brewer's yeast)
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MAGWDIFGWF RDVLASLGL W NKHGKLLF LG LDNAGKT TLL HMLKND RLAT LQPTW HPTSE ELAI GNIKFT TFD LGGHIQA RR LWKDYFPE V NGIVFLVDA ADPERFDEAR VELDALFNI A ELKDVPFV IL GNKIDAP NAV SEAELR SALG LLNTT ...String:
MAGWDIFGWF RDVLASLGL W NKHGKLLF LG LDNAGKT TLL HMLKND RLAT LQPTW HPTSE ELAI GNIKFT TFD LGGHIQA RR LWKDYFPE V NGIVFLVDA ADPERFDEAR VELDALFNI A ELKDVPFV IL GNKIDAP NAV SEAELR SALG LLNTT GSQRI EGQR PVEVFM CSV VMRNGYL EA FQWLSQYI

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Macromolecule #4: Sed5

MacromoleculeName: Sed5 / type: protein_or_peptide / ID: 4 / Enantiomer: LEVO
Source (natural)Organism: Saccharomyces cerevisiae (brewer's yeast)
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MNIKDRTSEF QQSVLSYKKR NKNFREQQRE RLQEKESENF ANNTTGNGKS VSEFQKKASG IAHEISSTAQ LLSKLAVLAK RKPMFNDNPV EIAELSFLIK RKIYAIEQSL VQLSQLKKTD VNGNTSNQSS KQPSAVQHSK NVVNLLNTQM KNISGSFKDV LEERQRLEMA ...String:
MNIKDRTSEF QQSVLSYKKR NKNFREQQRE RLQEKESENF ANNTTGNGKS VSEFQKKASG IAHEISSTAQ LLSKLAVLAK RKPMFNDNPV EIAELSFLIK RKIYAIEQSL VQLSQLKKTD VNGNTSNQSS KQPSAVQHSK NVVNLLNTQM KNISGSFKDV LEERQRLEMA NKDRWQKLTT DTGHAPADDQ TQSNHAADLT TYNNSNPFMT SLLDESSEKN NNSSNQGELS FPQNDSQLML MEEGQLSNNV YLQERNRAVE TIESTIQEVG NLFQQLASMV QEQGEVIQRI DANVDDIDLN ISGAQRELLK YFDRIKSNRG GGHHHHHH

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Experimental details

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Structure determination

Methodcryo EM
Processingsubtomogram averaging
Aggregation stateparticle

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Sample preparation

BufferpH: 6.8
GridModel: EMS Lacey Carbon / Material: COPPER / Pretreatment - Type: PLASMA CLEANING
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Specialist opticsEnergy filter - Slit width: 20 eV
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average electron dose: 3.46 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 3.5 µm / Nominal defocus min: 1.5 µm
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 4.13 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 4) / Number subtomograms used: 178700
ExtractionNumber tomograms: 326 / Number images used: 290000
Final angle assignmentType: MAXIMUM LIKELIHOOD
FSC plot (resolution estimation)

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