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- EMDB-15870: Subtomogram average of the human Sec61-TRAP-OSTA-translocon -

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Basic information

Entry
Database: EMDB / ID: EMD-15870
TitleSubtomogram average of the human Sec61-TRAP-OSTA-translocon
Map datasubtomogram average of the Sec61-TRAP-OSTA translocon
Sample
  • Organelle or cellular component: Sec61-TRAP-OSTA translocon complex
    • Protein or peptide: x 16 types
Function / homology
Function and homology information


oligosaccharyltransferase complex binding / oligosaccharyltransferase I complex / oligosaccharyltransferase III complex / Asparagine N-linked glycosylation / membrane-bounded organelle / endoplasmic reticulum Sec complex / co-translational protein modification / endoplasmic reticulum quality control compartment / pronephric nephron development / oligosaccharyltransferase complex ...oligosaccharyltransferase complex binding / oligosaccharyltransferase I complex / oligosaccharyltransferase III complex / Asparagine N-linked glycosylation / membrane-bounded organelle / endoplasmic reticulum Sec complex / co-translational protein modification / endoplasmic reticulum quality control compartment / pronephric nephron development / oligosaccharyltransferase complex / dolichyl-diphosphooligosaccharide-protein glycotransferase / dolichyl-diphosphooligosaccharide-protein glycotransferase activity / cotranslational protein targeting to membrane / Ssh1 translocon complex / Sec61 translocon complex / protein targeting to ER / protein insertion into ER membrane / protein N-linked glycosylation via asparagine / XBP1(S) activates chaperone genes / post-translational protein targeting to endoplasmic reticulum membrane / SRP-dependent cotranslational protein targeting to membrane, translocation / signal sequence binding / SRP-dependent cotranslational protein targeting to membrane / endoplasmic reticulum organization / post-translational protein targeting to membrane, translocation / protein N-linked glycosylation / epidermal growth factor binding / epithelial cell apoptotic process / retrograde protein transport, ER to cytosol / azurophil granule membrane / Insertion of tail-anchored proteins into the endoplasmic reticulum membrane / : / protein glycosylation / blastocyst development / Advanced glycosylation endproduct receptor signaling / SRP-dependent cotranslational protein targeting to membrane / protein transmembrane transporter activity / endomembrane system / enzyme activator activity / rough endoplasmic reticulum / T cell activation / response to endoplasmic reticulum stress / post-translational protein modification / response to cytokine / calcium channel activity / protein modification process / regulation of protein stability / melanosome / ribosome binding / ER-Phagosome pathway / Maturation of spike protein / membrane => GO:0016020 / nuclear body / inflammatory response / intracellular membrane-bounded organelle / apoptotic process / Neutrophil degranulation / positive regulation of cell population proliferation / endoplasmic reticulum membrane / negative regulation of apoptotic process / endoplasmic reticulum / protein-containing complex / RNA binding / extracellular exosome / membrane / metal ion binding / plasma membrane / cytosol / cytoplasm
Similarity search - Function
Translocon-associated protein (TRAP), alpha subunit / Translocon-associated protein (TRAP), alpha subunit / Translocon-associated / Translocon-associated protein subunit beta / Translocon-associated protein subunit gamma / Translocon-associated protein, delta subunit precursor (TRAP-delta) / Translocon-associated protein, gamma subunit (TRAP-gamma) / DAD/Ost2 / Oligosaccharyltransferase complex subunit / DAD family ...Translocon-associated protein (TRAP), alpha subunit / Translocon-associated protein (TRAP), alpha subunit / Translocon-associated / Translocon-associated protein subunit beta / Translocon-associated protein subunit gamma / Translocon-associated protein, delta subunit precursor (TRAP-delta) / Translocon-associated protein, gamma subunit (TRAP-gamma) / DAD/Ost2 / Oligosaccharyltransferase complex subunit / DAD family / Oligosaccharyltransferase subunit 5 / Dolichyl-diphosphooligosaccharide--protein glycosyltransferase 48kDa subunit / Oligosaccharyltransferase complex subunit OSTC / Oligosaccharyltransferase 48 kDa subunit beta / Dolichyl-diphosphooligosaccharide--protein glycosyltransferase subunit Swp1 / Oligosaccharyltransferase subunit Ribophorin II / Oligosaccaryltransferase / Oligosaccharyltransferase subunit ost4p superfamily / Oligosaccaryltransferase / Ribophorin I / Ribophorin I / Oligosaccharyl transferase complex, subunit OST3/OST6 / OST3 / OST6 family, transporter family / : / STT3/PglB/AglB core domain / : / Oligosaccharyl transferase, STT3 subunit / Oligosaccharyl transferase STT3, N-terminal / Protein transport Sec61-beta/Sbh / Protein transport protein SecG/Sec61-beta/Sbh / Sec61beta family / Protein translocase SEC61 complex, gamma subunit / Protein translocase SecE domain superfamily / Translocon Sec61/SecY, plug domain / Plug domain of Sec61p / Protein secE/sec61-gamma signature. / Protein secY signature 1. / Protein secY signature 2. / SecE/Sec61-gamma subunits of protein translocation complex / Protein translocase complex, SecE/Sec61-gamma subunit / SecY/SEC61-alpha family / SecY domain superfamily / SecY conserved site / SecY
Similarity search - Domain/homology
Dolichyl-diphosphooligosaccharide--protein glycosyltransferase subunit 1 / Dolichyl-diphosphooligosaccharide--protein glycosyltransferase subunit 2 / Dolichyl-diphosphooligosaccharide--protein glycosyltransferase subunit 4 / Dolichyl-diphosphooligosaccharide--protein glycosyltransferase 48 kDa subunit / Translocon-associated protein subunit alpha / Translocon-associated protein subunit beta / Dolichyl-diphosphooligosaccharide--protein glycosyltransferase subunit STT3A / Translocon-associated protein subunit delta / Protein transport protein Sec61 subunit gamma / Protein transport protein Sec61 subunit beta ...Dolichyl-diphosphooligosaccharide--protein glycosyltransferase subunit 1 / Dolichyl-diphosphooligosaccharide--protein glycosyltransferase subunit 2 / Dolichyl-diphosphooligosaccharide--protein glycosyltransferase subunit 4 / Dolichyl-diphosphooligosaccharide--protein glycosyltransferase 48 kDa subunit / Translocon-associated protein subunit alpha / Translocon-associated protein subunit beta / Dolichyl-diphosphooligosaccharide--protein glycosyltransferase subunit STT3A / Translocon-associated protein subunit delta / Protein transport protein Sec61 subunit gamma / Protein transport protein Sec61 subunit beta / Transmembrane protein 258 / Protein transport protein Sec61 subunit alpha isoform 1 / Dolichyl-diphosphooligosaccharide--protein glycosyltransferase subunit DAD1 / Oligosaccharyltransferase complex subunit OSTC / Translocon-associated protein subunit gamma
Similarity search - Component
Biological speciesHomo sapiens (human) / human (human)
Methodsubtomogram averaging / cryo EM / Resolution: 7.6 Å
AuthorsGemmer M / Fedry JMM / Forster FG
Funding supportEuropean Union, Netherlands, 3 items
OrganizationGrant numberCountry
European Research Council (ERC)724425European Union
Netherlands Organisation for Scientific Research (NWO)724.016.001 Netherlands
Netherlands Organisation for Scientific Research (NWO)212.152 Netherlands
CitationJournal: Nature / Year: 2023
Title: Visualization of translation and protein biogenesis at the ER membrane.
Authors: Max Gemmer / Marten L Chaillet / Joyce van Loenhout / Rodrigo Cuevas Arenas / Dimitrios Vismpas / Mariska Gröllers-Mulderij / Fujiet A Koh / Pascal Albanese / Richard A Scheltema / Stuart C ...Authors: Max Gemmer / Marten L Chaillet / Joyce van Loenhout / Rodrigo Cuevas Arenas / Dimitrios Vismpas / Mariska Gröllers-Mulderij / Fujiet A Koh / Pascal Albanese / Richard A Scheltema / Stuart C Howes / Abhay Kotecha / Juliette Fedry / Friedrich Förster /
Abstract: The dynamic ribosome-translocon complex, which resides at the endoplasmic reticulum (ER) membrane, produces a major fraction of the human proteome. It governs the synthesis, translocation, membrane ...The dynamic ribosome-translocon complex, which resides at the endoplasmic reticulum (ER) membrane, produces a major fraction of the human proteome. It governs the synthesis, translocation, membrane insertion, N-glycosylation, folding and disulfide-bond formation of nascent proteins. Although individual components of this machinery have been studied at high resolution in isolation, insights into their interplay in the native membrane remain limited. Here we use cryo-electron tomography, extensive classification and molecular modelling to capture snapshots of mRNA translation and protein maturation at the ER membrane at molecular resolution. We identify a highly abundant classical pre-translocation intermediate with eukaryotic elongation factor 1a (eEF1a) in an extended conformation, suggesting that eEF1a may remain associated with the ribosome after GTP hydrolysis during proofreading. At the ER membrane, distinct polysomes bind to different ER translocons specialized in the synthesis of proteins with signal peptides or multipass transmembrane proteins with the translocon-associated protein complex (TRAP) present in both. The near-complete atomic model of the most abundant ER translocon variant comprising the protein-conducting channel SEC61, TRAP and the oligosaccharyltransferase complex A (OSTA) reveals specific interactions of TRAP with other translocon components. We observe stoichiometric and sub-stoichiometric cofactors associated with OSTA, which are likely to include protein isomerases. In sum, we visualize ER-bound polysomes with their coordinated downstream machinery.
History
DepositionSep 27, 2022-
Header (metadata) releaseDec 7, 2022-
Map releaseDec 7, 2022-
UpdateFeb 8, 2023-
Current statusFeb 8, 2023Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_15870.png

Downloads & links

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Map

FileDownload / File: emd_15870.map.gz / Format: CCP4 / Size: 160.8 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotationsubtomogram average of the Sec61-TRAP-OSTA translocon
Projections & slices

Image control

Size
Brightness
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Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.72 Å/pix.
x 348 pix.
= 599.952 Å		1.72 Å/pix.
x 348 pix.
= 599.952 Å		1.72 Å/pix.
x 348 pix.
= 599.952 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.724 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.01
Minimum - Maximum-0.0659893 - 0.08555481
Average (Standard dev.)0.00024772485 (±0.0019177131)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions348348348
Spacing348348348
CellA=B=C: 599.95197 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: subtomogram average of the Sec61-TRAP-OSTA translocon

Fileemd_15870_half_map_1.map
Annotationsubtomogram average of the Sec61-TRAP-OSTA translocon
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: subtomogram average of the Sec61-TRAP-OSTA translocon

Fileemd_15870_half_map_2.map
Annotationsubtomogram average of the Sec61-TRAP-OSTA translocon
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Sec61-TRAP-OSTA translocon complex

EntireName: Sec61-TRAP-OSTA translocon complex
Components
  • Organelle or cellular component: Sec61-TRAP-OSTA translocon complex
    • Protein or peptide: Protein transport protein Sec61 subunit alpha isoform 1Protein targeting
    • Protein or peptide: Protein transport protein Sec61 subunit betaProtein targeting
    • Protein or peptide: Protein transport protein Sec61 subunit gammaProtein targeting
    • Protein or peptide: Signal peptide mix
    • Protein or peptide: Translocon-associated protein subunit alpha
    • Protein or peptide: Translocon-associated protein subunit beta
    • Protein or peptide: Translocon-associated protein subunit gamma
    • Protein or peptide: Translocon-associated protein subunit delta
    • Protein or peptide: Dolichyl-diphosphooligosaccharide--protein glycosyltransferase subunit STT3A
    • Protein or peptide: Oligosaccharyltransferase complex subunit OSTCOligosaccharyltransferase
    • Protein or peptide: Dolichyl-diphosphooligosaccharide--protein glycosyltransferase subunit 4
    • Protein or peptide: Transmembrane protein 258Transmembrane protein
    • Protein or peptide: Dolichyl-diphosphooligosaccharide--protein glycosyltransferase subunit DAD1
    • Protein or peptide: Dolichyl-diphosphooligosaccharide--protein glycosyltransferase 48 kDa subunit
    • Protein or peptide: Dolichyl-diphosphooligosaccharide--protein glycosyltransferase subunit 1
    • Protein or peptide: Dolichyl-diphosphooligosaccharide--protein glycosyltransferase subunit 2

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Supramolecule #1: Sec61-TRAP-OSTA translocon complex

SupramoleculeName: Sec61-TRAP-OSTA translocon complex / type: organelle_or_cellular_component / ID: 1 / Parent: 0 / Macromolecule list: all
Details: Translocon complex embedded in its native membrane and bound to cytosolic ribosomes
Source (natural)Organism: Homo sapiens (human)

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Macromolecule #1: Protein transport protein Sec61 subunit alpha isoform 1

MacromoleculeName: Protein transport protein Sec61 subunit alpha isoform 1
type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: human (human)
Molecular weightTheoretical: 52.304406 KDa
SequenceString: MAIKFLEVIK PFCVILPEIQ KPERKIQFKE KVLWTAITLF IFLVCCQIPL FGIMSSDSAD PFYWMRVILA SNRGTLMELG ISPIVTSGL IMQLLAGAKI IEVGDTPKDR ALFNGAQKLF GMIITIGQSI VYVMTGMYGD PSEMGAGICL LITIQLFVAG L IVLLLDEL ...String:
MAIKFLEVIK PFCVILPEIQ KPERKIQFKE KVLWTAITLF IFLVCCQIPL FGIMSSDSAD PFYWMRVILA SNRGTLMELG ISPIVTSGL IMQLLAGAKI IEVGDTPKDR ALFNGAQKLF GMIITIGQSI VYVMTGMYGD PSEMGAGICL LITIQLFVAG L IVLLLDEL LQKGYGLGSG ISLFIATNIC ETIVWKAFSP TTVNTGRGME FEGAIIALFH LLATRTDKVR ALREAFYRQN LP NLMNLIA TIFVFAVVIY FQGFRVDLPI KSARYRGQYN TYPIKLFYTS NIPIILQSAL VSNLYVISQM LSARFSGNLL VSL LGTWSD TSSGGPARAY PVGGLCYYLS PPESFGSVLE DPVHAVVYIV FMLGSCAFFS KTWIEVSGSS AKDVAKQLKE QQMV MRGHR ETSMVHELNR YIPTAAAFGG LCIGALSVLA DFLGAIGSGT GILLAVTIIY QYFEIFVKEQ SEVGSMGALL F

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Macromolecule #2: Protein transport protein Sec61 subunit beta

MacromoleculeName: Protein transport protein Sec61 subunit beta / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: human (human)
Molecular weightTheoretical: 9.987456 KDa
SequenceString:
MPGPTPSGTN VGSSGRSPSK AVAARAAGST VRQRKNASCG TRSAGRTTSA GTGGMWRFYT EDSPGLKVGP VPVLVMSLLF IASVFMLHI WGKYTRS

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Macromolecule #3: Protein transport protein Sec61 subunit gamma

MacromoleculeName: Protein transport protein Sec61 subunit gamma / type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: human (human)
Molecular weightTheoretical: 7.752325 KDa
SequenceString:
MDQVMQFVEP SRQFVKDSIR LVKRCTKPDR KEFQKIAMAT AIGFAIMGFI GFFVKLIHIP INNIIVGG

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Macromolecule #4: Signal peptide mix

MacromoleculeName: Signal peptide mix / type: protein_or_peptide / ID: 4 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 1.890321 KDa
SequenceString:
(UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK) (UNK)(UNK)(UNK)(UNK)(UNK)(UNK) (UNK)(UNK)(UNK) (UNK)(UNK)(UNK)

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Macromolecule #5: Translocon-associated protein subunit alpha

MacromoleculeName: Translocon-associated protein subunit alpha / type: protein_or_peptide / ID: 5 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: human (human)
Molecular weightTheoretical: 32.263219 KDa
SequenceString: MRLLPRLLLL LLLVFPATVL FRGGPRGLLA VAQDLTEDEE TVEDSIIEDE DDEAEVEEDE PTDLVEDKEE EDVSGEPEAS PSADTTILF VKGEDFPANN IVKFLVGFTN KGTEDFIVES LDASFRYPQD YQFYIQNFTA LPLNTVVPPQ RQATFEYSFI P AEPMGGRP ...String:
MRLLPRLLLL LLLVFPATVL FRGGPRGLLA VAQDLTEDEE TVEDSIIEDE DDEAEVEEDE PTDLVEDKEE EDVSGEPEAS PSADTTILF VKGEDFPANN IVKFLVGFTN KGTEDFIVES LDASFRYPQD YQFYIQNFTA LPLNTVVPPQ RQATFEYSFI P AEPMGGRP FGLVINLNYK DLNGNVFQDA VFNQTVTVIE REDGLDGETI FMYMFLAGLG LLVIVGLHQL LESRKRKRPI QK VEMGTSS QNDVDMSWIP QETLNQINKA SPRRLPRKRA QKRSVGSDE

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Macromolecule #6: Translocon-associated protein subunit beta

MacromoleculeName: Translocon-associated protein subunit beta / type: protein_or_peptide / ID: 6 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: human (human)
Molecular weightTheoretical: 20.154986 KDa
SequenceString:
MRLLSFVVLA LFAVTQAEEG ARLLASKSLL NRYAVEGRDL TLQYNIYNVG SSAALDVELS DDSFPPEDFG IVSGMLNVKW DRIAPASNV SHTVVLRPLK AGYFNFTSAT ITYLAQEDGP VVIGSTSAPG QGGILAQREF DRRFSPHFLD WAAFGVMTLP S IGIPLLLW YSSKRKYDTP KTKKN

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Macromolecule #7: Translocon-associated protein subunit gamma

MacromoleculeName: Translocon-associated protein subunit gamma / type: protein_or_peptide / ID: 7 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: human (human)
Molecular weightTheoretical: 21.106465 KDa
SequenceString:
MAPKGSSKQQ SEEDLLLQDF SRNLSAKSSA LFFGNAFIVS AIPIWLYWRI WHMDLIQSAV LYSVMTLVST YLVAFAYKNV KFVLKHKVA QKREDAVSKE VTRKLSEADN RKMSRKEKDE RILWKKNEVA DYEATTFSIF YNNTLFLVVV IVASFFILKN F NPTVNYIL SISASSGLIA LLSTGSK

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Macromolecule #8: Translocon-associated protein subunit delta

MacromoleculeName: Translocon-associated protein subunit delta / type: protein_or_peptide / ID: 8 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: human (human)
Molecular weightTheoretical: 19.015547 KDa
SequenceString:
MAAMASLGAL ALLLLSSLSR CSAEACLEPQ ITPSYYTTSD AVISTETVFI VEISLTCKNR VQNMALYADV GGKQFPVTRG QDVGRYQVS WSLDHKSAHA GTYEVRFFDE ESYSLLRKAQ RNNEDISIIP PLFTVSVDHR GTWNGPWVST EVLAAAIGLV I YYLAFSAK SHIQA

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Macromolecule #9: Dolichyl-diphosphooligosaccharide--protein glycosyltransferase su...

MacromoleculeName: Dolichyl-diphosphooligosaccharide--protein glycosyltransferase subunit STT3A
type: protein_or_peptide / ID: 9 / Number of copies: 1 / Enantiomer: LEVO
EC number: dolichyl-diphosphooligosaccharide-protein glycotransferase
Source (natural)Organism: human (human)
Molecular weightTheoretical: 80.60732 KDa
SequenceString: MTKFGFLRLS YEKQDTLLKL LILSMAAVLS FSTRLFAVLR FESVIHEFDP YFNYRTTRFL AEEGFYKFHN WFDDRAWYPL GRIIGGTIY PGLMITSAAI YHVLHFFHIT IDIRNVCVFL APLFSSFTTI VTYHLTKELK DAGAGLLAAA MIAVVPGYIS R SVAGSYDN ...String:
MTKFGFLRLS YEKQDTLLKL LILSMAAVLS FSTRLFAVLR FESVIHEFDP YFNYRTTRFL AEEGFYKFHN WFDDRAWYPL GRIIGGTIY PGLMITSAAI YHVLHFFHIT IDIRNVCVFL APLFSSFTTI VTYHLTKELK DAGAGLLAAA MIAVVPGYIS R SVAGSYDN EGIAIFCMLL TYYMWIKAVK TGSICWAAKC ALAYFYMVSS WGGYVFLINL IPLHVLVLML TGRFSHRIYV AY CTVYCLG TILSMQISFV GFQPVLSSEH MAAFGVFGLC QIHAFVDYLR SKLNPQQFEV LFRSVISLVG FVLLTVGALL MLT GKISPW TGRFYSLLDP SYAKNNIPII ASVSEHQPTT WSSYYFDLQL LVFMFPVGLY YCFSNLSDAR IFIIMYGVTS MYFS AVMVR LMLVLAPVMC ILSGIGVSQV LSTYMKNLDI SRPDKKSKKQ QDSTYPIKNE VASGMILVMA FFLITYTFHS TWVTS EAYS SPSIVLSARG GDGSRIIFDD FREAYYWLRH NTPEDAKVMS WWDYGYQITA MANRTILVDN NTWNNTHISR VGQAMA STE EKAYEIMREL DVSYVLVIFG GLTGYSSDDI NKFLWMVRIG GSTDTGKHIK ENDYYTPTGE FRVDREGSPV LLNCLMY KM CYYRFGQVYT EAKRPPGFDR VRNAEIGNKD FELDVLEEAY TTEHWLVRIY KVKDLDNRGL SRT

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Macromolecule #10: Oligosaccharyltransferase complex subunit OSTC

MacromoleculeName: Oligosaccharyltransferase complex subunit OSTC / type: protein_or_peptide / ID: 10 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: human (human)
Molecular weightTheoretical: 16.844215 KDa
SequenceString:
METLYRVPFL VLECPNLKLK KPPWLHMPSA MTVYALVVVS YFLITGGIIY DVIVEPPSVG SMTDEHGHQR PVAFLAYRVN GQYIMEGLA SSFLFTMGGL GFIILDRSNA PNIPKLNRFL LLFIGFVCVL LSFFMARVFM RMKLPGYLMG

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Macromolecule #11: Dolichyl-diphosphooligosaccharide--protein glycosyltransferase su...

MacromoleculeName: Dolichyl-diphosphooligosaccharide--protein glycosyltransferase subunit 4
type: protein_or_peptide / ID: 11 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: human (human)
Molecular weightTheoretical: 4.196004 KDa
SequenceString:
MITDVQLAIF ANMLGVSLFL LVVLYHYVAV NNPKKQE

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Macromolecule #12: Transmembrane protein 258

MacromoleculeName: Transmembrane protein 258 / type: protein_or_peptide / ID: 12 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: human (human)
Molecular weightTheoretical: 9.083804 KDa
SequenceString:
MELEAMSRYT SPVNPAVFPH LTVVLLAIGM FFTAWFFVYE VTSTKYTRDI YKELLISLVA SLFMGFGVLF LLLWVGIYV

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Macromolecule #13: Dolichyl-diphosphooligosaccharide--protein glycosyltransferase su...

MacromoleculeName: Dolichyl-diphosphooligosaccharide--protein glycosyltransferase subunit DAD1
type: protein_or_peptide / ID: 13 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: human (human)
Molecular weightTheoretical: 12.503631 KDa
SequenceString:
MSASVVSVIS RFLEEYLSST PQRLKLLDAY LLYILLTGAL QFGYCLLVGT FPFNSFLSGF ISCVGSFILA VCLRIQINPQ NKADFQGIS PERAFADFLF ASTILHLVVM NFVG

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Macromolecule #14: Dolichyl-diphosphooligosaccharide--protein glycosyltransferase 48...

MacromoleculeName: Dolichyl-diphosphooligosaccharide--protein glycosyltransferase 48 kDa subunit
type: protein_or_peptide / ID: 14 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: human (human)
Molecular weightTheoretical: 50.854578 KDa
SequenceString: MGYFRCARAG SFGRRRKMEP STAARAWALF WLLLPLLGAV CASGPRTLVL LDNLNVRETH SLFFRSLKDR GFELTFKTAD DPSLSLIKY GEFLYDNLII FSPSVEDFGG NINVETISAF IDGGGSVLVA ASSDIGDPLR ELGSECGIEF DEEKTAVIDH H NYDISDLG ...String:
MGYFRCARAG SFGRRRKMEP STAARAWALF WLLLPLLGAV CASGPRTLVL LDNLNVRETH SLFFRSLKDR GFELTFKTAD DPSLSLIKY GEFLYDNLII FSPSVEDFGG NINVETISAF IDGGGSVLVA ASSDIGDPLR ELGSECGIEF DEEKTAVIDH H NYDISDLG QHTLIVADTE NLLKAPTIVG KSSLNPILFR GVGMVADPDN PLVLDILTGS STSYSFFPDK PITQYPHAVG KN TLLIAGL QARNNARVIF SGSLDFFSDS FFNSAVQKAA PGSQRYSQTG NYELAVALSR WVFKEEGVLR VGPVSHHRVG ETA PPNAYT VTDLVEYSIV IQQLSNGKWV PFDGDDIQLE FVRIDPFVRT FLKKKGGKYS VQFKLPDVYG VFQFKVDYNR LGYT HLYSS TQVSVRPLQH TQYERFIPSA YPYYASAFSM MLGLFIFSIV FLHMKEKEKS D

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Macromolecule #15: Dolichyl-diphosphooligosaccharide--protein glycosyltransferase su...

MacromoleculeName: Dolichyl-diphosphooligosaccharide--protein glycosyltransferase subunit 1
type: protein_or_peptide / ID: 15 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: human (human)
Molecular weightTheoretical: 68.656156 KDa
SequenceString: MEAPAAGLFL LLLLGTWAPA PGSASSEAPP LINEDVKRTV DLSSHLAKVT AEVVLAHLGG GSTSRATSFL LALEPELEAR LAHLGVQVK GEDEEENNLE VRETKIKGKS GRFFTVKLPV ALDPGAKISV IVETVYTHVL HPYPTQITQS EKQFVVFEGN H YFYSPYPT ...String:
MEAPAAGLFL LLLLGTWAPA PGSASSEAPP LINEDVKRTV DLSSHLAKVT AEVVLAHLGG GSTSRATSFL LALEPELEAR LAHLGVQVK GEDEEENNLE VRETKIKGKS GRFFTVKLPV ALDPGAKISV IVETVYTHVL HPYPTQITQS EKQFVVFEGN H YFYSPYPT KTQTMRVKLA SRNVESYTKL GNPTRSEDLL DYGPFRDVPA YSQDTFKVHY ENNSPFLTIT SMTRVIEVSH WG NIAVEEN VDLKHTGAVL KGPFSRYDYQ RQPDSGISSI RSFKTILPAA AQDVYYRDEI GNVSTSHLLI LDDSVEMEIR PRF PLFGGW KTHYIVGYNL PSYEYLYNLG DQYALKMRFV DHVFDEQVID SLTVKIILPE GAKNIEIDSP YEISRAPDEL HYTY LDTFG RPVIVAYKKN LVEQHIQDIV VHYTFNKVLM LQEPLLVVAA FYILFFTVII YVRLDFSITK DPAAEARMKV ACITE QVLT LVNKRIGLYR HFDETVNRYK QSRDISTLNS GKKSLETEHK ALTSEIALLQ SRLKTEGSDL CDRVSEMQKL DAQVKE LVL KSAVEAERLV AGKLKKDTYI ENEKLISGKR QELVTKIDHI LDAL

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Macromolecule #16: Dolichyl-diphosphooligosaccharide--protein glycosyltransferase su...

MacromoleculeName: Dolichyl-diphosphooligosaccharide--protein glycosyltransferase subunit 2
type: protein_or_peptide / ID: 16 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: human (human)
Molecular weightTheoretical: 69.347508 KDa
SequenceString: MAPPGSSTVF LLALTIIAST WALTPTHYLT KHDVERLKAS LDRPFTNLES AFYSIVGLSS LGAQVPDAKK ACTYIRSNLD PSNVDSLFY AAQASQALSG CEISISNETK DLLLAAVSED SSVTQIYHAV AALSGFGLPL ASQEALSALT ARLSKEETVL A TVQALQTA ...String:
MAPPGSSTVF LLALTIIAST WALTPTHYLT KHDVERLKAS LDRPFTNLES AFYSIVGLSS LGAQVPDAKK ACTYIRSNLD PSNVDSLFY AAQASQALSG CEISISNETK DLLLAAVSED SSVTQIYHAV AALSGFGLPL ASQEALSALT ARLSKEETVL A TVQALQTA SHLSQQADLR SIVEEIEDLV ARLDELGGVY LQFEEGLETT ALFVAATYKL MDHVGTEPSI KEDQVIQLMN AI FSKKNFE SLSEAFSVAS AAAVLSHNRY HVPVVVVPEG SASDTHEQAI LRLQVTNVLS QPLTQATVKL EHAKSVASRA TVL QKTSFT PVGDVFELNF MNVKFSSGYY DFLVEVEGDN RYIANTVELR VKISTEVGIT NVDLSTVDKD QSIAPKTTRV TYPA KAKGT FIADSHQNFA LFFQLVDVNT GAELTPHQTF VRLHNQKTGQ EVVFVAEPDN KNVYKFELDT SERKIEFDSA SGTYT LYLI IGDATLKNPI LWNVADVVIK FPEEEAPSTV LSQNLFTPKQ EIQHLFREPE KRPPTVVSNT FTALILSPLL LLFALW IRI GANVSNFTFA PSTIIFHLGH AAMLGLMYVY WTQLNMFQTL KYLAILGSVT FLAGNRMLAQ QAVKRTAH

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Experimental details

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Structure determination

Methodcryo EM
Processingsubtomogram averaging
Aggregation stateparticle

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Sample preparation

Concentration2 mg/mL
BufferpH: 7.4
GridModel: Quantifoil / Material: COPPER / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 30 sec.
VitrificationCryogen name: ETHANE-PROPANE / Details: Manual plunger.
DetailsER-derived vesicles decorated with cytosolic ribosomes.

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Electron microscopy

MicroscopeFEI TALOS ARCTICA
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 100.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 3.0 µm / Nominal defocus min: 3.0 µm / Nominal magnification: 79000
Sample stageCooling holder cryogen: NITROGEN
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Digitization - Frames/image: 7-7 / Number grids imaged: 8 / Average exposure time: 0.7 sec. / Average electron dose: 2.3 e/Å2
Experimental equipment
Model: Talos Arctica / Image courtesy: FEI Company

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Image processing

ExtractionNumber tomograms: 869 / Number images used: 134350
Reference model: Subtomogram average of ER membrane-associated ribosome
Method: Template matching / Software - Name: PyTom (ver. 0.994)
Final 3D classificationSoftware - Name: RELION (ver. 3.1.1)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.1.1)
Final reconstructionAlgorithm: BACK PROJECTION / Resolution.type: BY AUTHOR / Resolution: 7.6 Å / Resolution method: FSC 0.143 CUT-OFF / Number subtomograms used: 42215

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Atomic model buiding 1

RefinementSpace: REAL / Protocol: FLEXIBLE FIT
Output model

PDB-8b6l:
Subtomogram average of the human Sec61-TRAP-OSTA-translocon

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