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- EMDB-32822: An apo TRiC map -

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Basic information

Entry
Database: EMDB / ID: EMD-32822
TitleAn apo TRiC map
Map dataA sharpened map of apo TRiC
Sample
  • Complex: Apo TRiC/CCT
    • Other: CCT subunit 1
    • Other: CCT subunit 2
    • Other: CCT subunit 3
    • Other: CCT subunit 4
    • Other: CCT subunit 5
    • Other: CCT subunit 6
    • Other: CCT subunit 7
    • Other: CCT subunit 8
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.11 Å
AuthorsPark JS / Roh SH
Funding support Korea, Republic Of, 1 items
OrganizationGrant numberCountry
National Research Foundation (NRF, Korea) Korea, Republic Of
CitationJournal: Cell / Year: 2022
Title: Structural visualization of the tubulin folding pathway directed by human chaperonin TRiC/CCT.
Authors: Daniel Gestaut / Yanyan Zhao / Junsun Park / Boxue Ma / Alexander Leitner / Miranda Collier / Grigore Pintilie / Soung-Hun Roh / Wah Chiu / Judith Frydman /
Abstract: The ATP-dependent ring-shaped chaperonin TRiC/CCT is essential for cellular proteostasis. To uncover why some eukaryotic proteins can only fold with TRiC assistance, we reconstituted the folding of ...The ATP-dependent ring-shaped chaperonin TRiC/CCT is essential for cellular proteostasis. To uncover why some eukaryotic proteins can only fold with TRiC assistance, we reconstituted the folding of β-tubulin using human prefoldin and TRiC. We find unstructured β-tubulin is delivered by prefoldin to the open TRiC chamber followed by ATP-dependent chamber closure. Cryo-EM resolves four near-atomic-resolution structures containing progressively folded β-tubulin intermediates within the closed TRiC chamber, culminating in native tubulin. This substrate folding pathway appears closely guided by site-specific interactions with conserved regions in the TRiC chamber. Initial electrostatic interactions between the TRiC interior wall and both the folded tubulin N domain and its C-terminal E-hook tail establish the native substrate topology, thus enabling C-domain folding. Intrinsically disordered CCT C termini within the chamber promote subsequent folding of tubulin's core and middle domains and GTP-binding. Thus, TRiC's chamber provides chemical and topological directives that shape the folding landscape of its obligate substrates.
History
DepositionFeb 7, 2022-
Header (metadata) releaseJan 11, 2023-
Map releaseJan 11, 2023-
UpdateJan 11, 2023-
Current statusJan 11, 2023Processing site: PDBj / Status: Released

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Structure visualization

Supplemental images

emd_32822.png

Downloads & links

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Map

FileDownload / File: emd_32822.map.gz / Format: CCP4 / Size: 125 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationA sharpened map of apo TRiC
Voxel sizeX=Y=Z: 1 Å
Density
Contour LevelBy AUTHOR: 0.67
Minimum - Maximum-5.5629873 - 9.041815
Average (Standard dev.)0.008324801 (±0.24187607)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions320320320
Spacing320320320
CellA=B=C: 320.0 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: A half map of apo TRiC

Fileemd_32822_half_map_1.map
AnnotationA half map of apo TRiC
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: A half map of apo TRiC

Fileemd_32822_half_map_2.map
AnnotationA half map of apo TRiC
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Apo TRiC/CCT

EntireName: Apo TRiC/CCT
Components
  • Complex: Apo TRiC/CCT
    • Other: CCT subunit 1
    • Other: CCT subunit 2
    • Other: CCT subunit 3
    • Other: CCT subunit 4
    • Other: CCT subunit 5
    • Other: CCT subunit 6
    • Other: CCT subunit 7
    • Other: CCT subunit 8

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Supramolecule #1: Apo TRiC/CCT

SupramoleculeName: Apo TRiC/CCT / type: complex / ID: 1 / Chimera: Yes / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 1 MDa

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Macromolecule #1: CCT subunit 1

MacromoleculeName: CCT subunit 1 / type: other / ID: 1 / Classification: other
Source (natural)Organism: Homo sapiens (human)
SequenceString: MEGPLSVFGD RSTGETIRSQ NVMAAASIAN IVKSSLGPVG LDKMLVDDIG DVTITNDGAT ILKLLEVEH PAAKVLCELA DLQDKEVGDG TTSVVIIAAE LLKNADELVK QKIHPTSVIS G YRLACKEA VRYINENLIV NTDELGRDCL INAAKTSMSS KIIGINGDFF ...String:
MEGPLSVFGD RSTGETIRSQ NVMAAASIAN IVKSSLGPVG LDKMLVDDIG DVTITNDGAT ILKLLEVEH PAAKVLCELA DLQDKEVGDG TTSVVIIAAE LLKNADELVK QKIHPTSVIS G YRLACKEA VRYINENLIV NTDELGRDCL INAAKTSMSS KIIGINGDFF ANMVVDAVLA IK YTDIRGQ PRYPVNSVNI LKAHGRSQME SMLISGYALN CVVGSQGMPK RIVNAKIACL DFS LQKTKM KLGVQVVITD PEKLDQIRQR ESDITKERIQ KILATGANVI LTTGGIDDMC LKYF VEAGA MAVRRVLKRD LKRIAKASGA TILSTLANLE GEETFEAAML GQAEEVVQER ICDDE LILI KNTKARTSAS IILRGANDFM CDEMERSLHD ALCVVKRVLE SKSVVPGGGA VEAALS IYL ENYATSMGSR EQLAIAEFAR SLLVIPNTLA VNAAQDSTDL VAKLRAFHNE AQVNPER KN LKWIGLDLSN GKPRDNKQAG VFEPTIVKVK SLKFATEAAI TILRIDDLIK LHPESKDD K HGSYEDAVHS GALND
Recombinant expressionOrganism: Trichoplusia ni (cabbage looper)

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Macromolecule #2: CCT subunit 2

MacromoleculeName: CCT subunit 2 / type: other / ID: 2 / Classification: other
Source (natural)Organism: Homo sapiens (human)
SequenceString: MASLSLAPVN IFKAGADEER AETARLTSFI GAIAIGDLVK STLGPKGMDK ILLSSGRDAS LMVTNDGAT ILKNIGVDNP AAKVLVDMSR VQDDEVGDGT TSVTVLAAEL LREAESLIAK K IHPQTIIA GWREATKAAR EALLSSAVDH GSDEVKFRQD LMNIAGTTLS ...String:
MASLSLAPVN IFKAGADEER AETARLTSFI GAIAIGDLVK STLGPKGMDK ILLSSGRDAS LMVTNDGAT ILKNIGVDNP AAKVLVDMSR VQDDEVGDGT TSVTVLAAEL LREAESLIAK K IHPQTIIA GWREATKAAR EALLSSAVDH GSDEVKFRQD LMNIAGTTLS SKLLTHHKDH FT KLAVEAV LRLKGSGNLE AIHIIKKLGG SLADSYLDEG FLLDKKIGVN QPKRIENAKI LIA NTGMDT DKIKIFGSRV RVDSTAKVAE IEHAEKEKMK EKVERILKHG INCFINRQLI YNYP EQLFG AAGVMAIEHA DFAGVERLAL VTGGEIASTF DHPELVKLGS CKLIEEVMIG EDKLI HFSG VALGEACTIV LRGATQQILD EAERSLHDAL CVLAQTVKDS RTVYGGGCSE MLMAHA VTQ LANRTPGKEA VAMESYAKAL RMLPTIIADN AGYDSADLVA QLRAAHSEGN TTAGLDM RE GTIGDMAILG ITESFQVKRQ VLLSAAEAAE VILRVDNIIK AAPRKRVPDH HPC
Recombinant expressionOrganism: Trichoplusia ni (cabbage looper)

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Macromolecule #3: CCT subunit 3

MacromoleculeName: CCT subunit 3 / type: other / ID: 3 / Classification: other
Source (natural)Organism: Homo sapiens (human)
SequenceString: MMGHRPVLVL SQNTKRESGR KVQSGNINAA KTIADIIRTC LGPKSMMKML LDPMGGIVMT NDGNAILRE IQVQHPAAKS MIEISRTQDE EVGDGTTSVI ILAGEMLSVA EHFLEQQMHP T VVISAYRK ALDDMISTLK KISIPVDISD SDMMLNIINS SITTKAISRW ...String:
MMGHRPVLVL SQNTKRESGR KVQSGNINAA KTIADIIRTC LGPKSMMKML LDPMGGIVMT NDGNAILRE IQVQHPAAKS MIEISRTQDE EVGDGTTSVI ILAGEMLSVA EHFLEQQMHP T VVISAYRK ALDDMISTLK KISIPVDISD SDMMLNIINS SITTKAISRW SSLACNIALD AV KMVQFEE NGRKEIDIKK YARVEKIPGG IIEDSCVLRG VMINKDVTHP RMRRYIKNPR IVL LDSSLE YKKGESQTDI EITREEDFTR ILQMEEEYIQ QLCEDIIQLK PDVVITEKGI SDLA QHYLM RANITAIRRV RKTDNNRIAR ACGARIVSRP EELREDDVGT GAGLLEIKKI GDEYF TFIT DCKDPKACTI LLRGASKEIL SEVERNLQDA MQVCRNVLLD PQLVPGGGAS EMAVAH ALT EKSKAMTGVE QWPYRAVAQA LEVIPRTLIQ NCGASTIRLL TSLRAKHTQE NCETWGV NG ETGTLVDMKE LGIWEPLAVK LQTYKTAVET AVLLLRIDDI VSGHKKKGDD QSRQGGAP D AGQE
Recombinant expressionOrganism: Trichoplusia ni (cabbage looper)

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Macromolecule #4: CCT subunit 4

MacromoleculeName: CCT subunit 4 / type: other / ID: 4 / Classification: other
Source (natural)Organism: Homo sapiens (human)
SequenceString: MPENVAPRSG ATAGAAGGRG KGAYQDRDKP AQIRFSNISA AKAVADAIRT SLGPKGMDKM IQDGKGDVT ITNDGATILK QMQVLHPAAR MLVELSKAQD IEAGDGTTSV VIIAGSLLDS C TKLLQKGI HPTIISESFQ KALEKGIEIL TDMSRPVELS DRETLLNSAT ...String:
MPENVAPRSG ATAGAAGGRG KGAYQDRDKP AQIRFSNISA AKAVADAIRT SLGPKGMDKM IQDGKGDVT ITNDGATILK QMQVLHPAAR MLVELSKAQD IEAGDGTTSV VIIAGSLLDS C TKLLQKGI HPTIISESFQ KALEKGIEIL TDMSRPVELS DRETLLNSAT TSLNSKVVSQ YS SLLSPMS VNAVMKVIDP ATATSVDLRD IKIVKKLGGT IDDCELVEGL VLTQKVSNSG ITR VEKAKI GLIQFCLSAP KTDMDNQIVV SDYAQMDRVL REERAYILNL VKQIKKTGCN VLLI QKSIL RDALSDLALH FLNKMKIMVI KDIEREDIEF ICKTIGTKPV AHIDQFTADM LGSAE LAEE VNLNGSGKLL KITGCASPGK TVTIVVRGSN KLVIEEAERS IHDALCVIRC LVKKRA LIA GGGAPEIELA LRLTEYSRTL SGMESYCVRA FADAMEVIPS TLAENAGLNP ISTVTEL RN RHAQGEKTAG INVRKGGISN ILEELVVQPL LVSVSALTLA TETVRSILKI DDVVNTR
Recombinant expressionOrganism: Trichoplusia ni (cabbage looper)

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Macromolecule #5: CCT subunit 5

MacromoleculeName: CCT subunit 5 / type: other / ID: 5 / Classification: other
Source (natural)Organism: Homo sapiens (human)
SequenceString: MASMGTLAFD EYGRPFLIIK DQDRKSRLMG LEALKSHIMA AKAVANTMRT SLGPNGLDKM MVDKDGDVT VTNDGATILS MMDVDHQIAK LMVELSKSQD DEIGDGTTGV VVLAGALLEE A EQLLDRGI HPIRIADGYE QAARVAIEHL DKISDSVLVD IKDTEPLIQT ...String:
MASMGTLAFD EYGRPFLIIK DQDRKSRLMG LEALKSHIMA AKAVANTMRT SLGPNGLDKM MVDKDGDVT VTNDGATILS MMDVDHQIAK LMVELSKSQD DEIGDGTTGV VVLAGALLEE A EQLLDRGI HPIRIADGYE QAARVAIEHL DKISDSVLVD IKDTEPLIQT AKTTLGSKVV NS CHRQMAE IAVNAVLTVA DMERRDVDFE LIKVEGKVGG RLEDTKLIKG VIVDKDFSHP QMP KKVEDA KIAILTCPFE PPKPKTKHKL DVTSVEDYKA LQKYEKEKFE EMIQQIKETG ANLA ICQWG FDDEANHLLL QNNLPAVRWV GGPEIELIAI ATGGRIVPRF SELTAEKLGF AGLVQ EISF GTTKDKMLVI EQCKNSRAVT IFIRGGNKMI IEEAKRSLHD ALCVIRNLIR DNRVVY GGG AAEISCALAV SQEADKCPTL EQYAMRAFAD ALEVIPMALS ENSGMNPIQT MTEVRAR QV KEMNPALGID CLHKGTNDMK QQHVIETLIG KKQQISLATQ MVRMILKIDD IRKPGESEE
Recombinant expressionOrganism: Trichoplusia ni (cabbage looper)

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Macromolecule #6: CCT subunit 6

MacromoleculeName: CCT subunit 6 / type: other / ID: 6 / Classification: other
Source (natural)Organism: Homo sapiens (human)
SequenceString: MAAVKTLNPK AEVARAQAAL AVNISAARGL QDVLRTNLGP KGTMKMLVSG AGDIKLTKDG NVLLHEMQI QHPTASLIAK VATAQDDITG DGTTSNVLII GELLKQADLY ISEGLHPRII T EGFEAAKE KALQFLEEVK VSREMDRETL IDVARTSLRT KVHAELADVL ...String:
MAAVKTLNPK AEVARAQAAL AVNISAARGL QDVLRTNLGP KGTMKMLVSG AGDIKLTKDG NVLLHEMQI QHPTASLIAK VATAQDDITG DGTTSNVLII GELLKQADLY ISEGLHPRII T EGFEAAKE KALQFLEEVK VSREMDRETL IDVARTSLRT KVHAELADVL TEAVVDSILA IK KQDEPID LFMIEIMEMK HKSETDTSLI RGLVLDHGAR HPDMKKRVED AYILTCNVSL EYE KTEVNS GFFYKSAEER EKLVKAERKF IEDRVKKIIE LKRKVCGDSD KGFVVINQKG IDPF SLDAL SKEGIVALRR AKRRNMERLT LACGGVALNS FDDLSPDCLG HAGLVYEYTL GEEKF TFIE KCNNPRSVTL LIKGPNKHTL TQIKDAVRDG LRAVKNAIDD GCVVPGAGAV EVAMAE ALI KHKPSVKGRA QLGVQAFADA LLIIPKVLAQ NSGFDLQETL VKIQAEHSES GQLVGVD LN TGEPMVAAEV GVWDNYCVKK QLLHSCTVIA TNILLVDEIM RAGMSFLKG
Recombinant expressionOrganism: Trichoplusia ni (cabbage looper)

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Macromolecule #7: CCT subunit 7

MacromoleculeName: CCT subunit 7 / type: other / ID: 7 / Classification: other
Source (natural)Organism: Homo sapiens (human)
SequenceString: MMPTPVILLK EGTDSSQGIP QLVSNISACQ VIAEAVRTTL GPRGMDKLIV DGRGKATISN DGATILKLL DVVHPAAKTL VDIAKSQDAE VGDGTTSVTL LAAEFLKQVK PYVEEGLHPQ I IIRAFRTA TQLAVNKIKE IAVTVKKADK VEQRKLLEKC AMTALSSKLI ...String:
MMPTPVILLK EGTDSSQGIP QLVSNISACQ VIAEAVRTTL GPRGMDKLIV DGRGKATISN DGATILKLL DVVHPAAKTL VDIAKSQDAE VGDGTTSVTL LAAEFLKQVK PYVEEGLHPQ I IIRAFRTA TQLAVNKIKE IAVTVKKADK VEQRKLLEKC AMTALSSKLI SQQKAFFAKM VV DAVMMLD DLLQLKMIGI KKVQGGALED SQLVAGVAFK KTFSYAGFEM QPKKYHNPKI ALL NVELEL KAEKDNAEIR VHTVEDYQAI VDAEWNILYD KLEKIHHSGA KVVLSKLPIG DVAT QYFAD RDMFCAGRVP EEDLKRTMMA CGGSIQTSVN ALSADVLGRC QVFEETQIGG ERYNF FTGC PKAKTCTFIL RGGAEQFMEE TERSLHDAIM IVRRAIKNDS VVAGGGAIEM ELSKYL RDY SRTIPGKQQL LIGAYAKALE IIPRQLCDNA GFDATNILNK LRARHAQGGS HHHHHHG SG TWYGVDINNE DIADNFEAFV WEPAMVRINA LTAASEAACL IVSVDETIKN PRSTVDAP T AAGRGRGRGR PH
Recombinant expressionOrganism: Trichoplusia ni (cabbage looper)

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Macromolecule #8: CCT subunit 8

MacromoleculeName: CCT subunit 8 / type: other / ID: 8 / Classification: other
Source (natural)Organism: Homo sapiens (human)
SequenceString: MALHVPKAPG FAQMLKEGAK HFSGLEEAVY RNIQACKELA QTTRTAYGPN GMNKMVINHL EKLFVTNDA ATILRELEVQ HPAAKMIVMA SHMQEQEVGD GTNFVLVFAG ALLELAEELL R IGLSVSEV IEGYEIACRK AHEILPNLVC CSAKNLRDID EVSSLLRTSI ...String:
MALHVPKAPG FAQMLKEGAK HFSGLEEAVY RNIQACKELA QTTRTAYGPN GMNKMVINHL EKLFVTNDA ATILRELEVQ HPAAKMIVMA SHMQEQEVGD GTNFVLVFAG ALLELAEELL R IGLSVSEV IEGYEIACRK AHEILPNLVC CSAKNLRDID EVSSLLRTSI MSKQYGNEVF LA KLIAQAC VSIFPDSGHF NVDNIRVCKI LGSGISSSSV LHGMVFKKET EGDVTSVKDA KIA VYSCPF DGMITETKGT VLIKTAEELM NFSKGEENLM DAQVKAIADT GANVVVTGGK VADM ALHYA NKYNIMLVRL NSKWDLRRLC KTVGATALPR LTPPVLEEMG HCDSVYLSEV GDTQV VVFK HEKEDGAIST IVLRGSTDNL MDDIERAVDD GVNTFKVLTR DKRLVPGGGA TEIELA KQI TSYGETCPGL EQYAIKKFAE AFEAIPRALA ENSGVKANEV ISKLYAVHQE GNKNVGL DI EAEVPAVKDM LEAGILDTYL GKYWAIKLAT NAAVTVLRVD QIIMAKPAGG PKPPSGKK D WDDDQN
Recombinant expressionOrganism: Trichoplusia ni (cabbage looper)

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation state3D array

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Sample preparation

BufferpH: 7.4
Component:
ConcentrationFormulaName
50.0 mMNaClSodium chloridesodium chloride
1.0 mMC4H10O2S2DTT
GridModel: Quantifoil R1.2/1.3 / Support film - Material: CARBON / Support film - topology: HOLEY
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 3.5 µm / Nominal defocus min: 0.5 µm
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Number real images: 6084 / Average electron dose: 50.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 945248
Details: The initial particle selection after 2D class classification
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 3.2.0)
Final 3D classificationNumber classes: 4 / Avg.num./class: 300000 / Software - Name: cryoSPARC (ver. 3.2.0) / Details: The final 3D classification
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 3.2.0)
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.11 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 3.2.0) / Number images used: 662744
FSC plot (resolution estimation)

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Atomic model buiding 1

RefinementSpace: REAL / Protocol: RIGID BODY FIT

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