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- EMDB-24884: apo NEXT overall reconstruction -

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Open data


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Basic information

Entry
Database: EMDB / ID: EMD-24884
Titleapo NEXT overall reconstruction
Map dataapo NEXT overall map
Sample
  • Complex: human Nuclear Exosome Targeting (NEXT) complex
    • Protein or peptide: MTR4
    • Protein or peptide: ZCCHC8
    • Protein or peptide: RBM7
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 8.36 Å
AuthorsPuno MR / Lima CD
Funding support United States, 2 items
OrganizationGrant numberCountry
Howard Hughes Medical Institute (HHMI) United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM118080 United States
CitationJournal: Cell / Year: 2022
Title: Structural basis for RNA surveillance by the human nuclear exosome targeting (NEXT) complex.
Authors: M Rhyan Puno / Christopher D Lima /
Abstract: RNA quality control relies on co-factors and adaptors to identify and prepare substrates for degradation by ribonucleases such as the 3' to 5' ribonucleolytic RNA exosome. Here, we determined ...RNA quality control relies on co-factors and adaptors to identify and prepare substrates for degradation by ribonucleases such as the 3' to 5' ribonucleolytic RNA exosome. Here, we determined cryogenic electron microscopy structures of human nuclear exosome targeting (NEXT) complexes bound to RNA that reveal mechanistic insights to substrate recognition and early steps that precede RNA handover to the exosome. The structures illuminate ZCCHC8 as a scaffold, mediating homodimerization while embracing the MTR4 helicase and flexibly anchoring RBM7 to the helicase core. All three subunits collaborate to bind the RNA, with RBM7 and ZCCHC8 surveying sequences upstream of the 3' end to facilitate RNA capture by MTR4. ZCCHC8 obscures MTR4 surfaces important for RNA binding and extrusion as well as MPP6-dependent recruitment and docking onto the RNA exosome core, interactions that contribute to RNA surveillance by coordinating RNA capture, translocation, and extrusion from the helicase to the exosome for decay.
History
DepositionSep 15, 2021-
Header (metadata) releaseJun 15, 2022-
Map releaseJun 15, 2022-
UpdateJun 22, 2022-
Current statusJun 22, 2022Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_24884.png

Downloads & links

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Map

FileDownload / File: emd_24884.map.gz / Format: CCP4 / Size: 216 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotationapo NEXT overall map
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.09 Å/pix.
x 384 pix.
= 417.792 Å		1.09 Å/pix.
x 384 pix.
= 417.792 Å		1.09 Å/pix.
x 384 pix.
= 417.792 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.088 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.008
Minimum - Maximum-0.008346002 - 0.03188567
Average (Standard dev.)7.062381e-05 (±0.0009916171)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions384384384
Spacing384384384
CellA=B=C: 417.79202 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: half map 1

Fileemd_24884_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 1

Fileemd_24884_half_map_2.map
Annotationhalf map 1
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : human Nuclear Exosome Targeting (NEXT) complex

EntireName: human Nuclear Exosome Targeting (NEXT) complex
Components
  • Complex: human Nuclear Exosome Targeting (NEXT) complex
    • Protein or peptide: MTR4
    • Protein or peptide: ZCCHC8
    • Protein or peptide: RBM7

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Supramolecule #1: human Nuclear Exosome Targeting (NEXT) complex

SupramoleculeName: human Nuclear Exosome Targeting (NEXT) complex / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Homo sapiens (human)
Recombinant expressionOrganism: Escherichia coli (E. coli)

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Macromolecule #1: MTR4

MacromoleculeName: MTR4 / type: protein_or_peptide / ID: 1
Details: The first three amino acid residues (SGD) in the sample sequence are cloning artefact.
Enantiomer: LEVO / EC number: RNA helicase
Source (natural)Organism: Homo sapiens (human)
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: SGDMADAFGD ELFSVFEGDS TTAAGTKKDK EKDKGKWKGP PGSADKAGKR FDGKLQSEST NNGKNKRDVD FEGTDEPIFG KKPRIEESI TEDLSLADLM PRVKVQSVET VEGCTHEVAL PAEEDYLPLK PRVGKAAKEY PFILDAFQRE AIQCVDNNQS V LVSAHTSA ...String:
SGDMADAFGD ELFSVFEGDS TTAAGTKKDK EKDKGKWKGP PGSADKAGKR FDGKLQSEST NNGKNKRDVD FEGTDEPIFG KKPRIEESI TEDLSLADLM PRVKVQSVET VEGCTHEVAL PAEEDYLPLK PRVGKAAKEY PFILDAFQRE AIQCVDNNQS V LVSAHTSA GKTVCAEYAI ALALREKQRV IFTSPIKALS NQKYREMYEE FQDVGLMTGD VTINPTASCL VMTTEILRSM LY RGSEVMR EVAWVIFDEI HYMRDSERGV VWEETIILLP DNVHYVFLSA TIPNARQFAE WICHLHKQPC HVIYTDYRPT PLQ HYIFPA GGDGLHLVVD ENGDFREDNF NTAMQVLRDA GDLAKGDQKG RKGGTKGPSN VFKIVKMIME RNFQPVIIFS FSKK DCEAY ALQMTKLDFN TDEEKKMVEE VFSNAIDCLS DEDKKLPQVE HVLPLLKRGI GIHHGGLLPI LKETIEILFS EGLIK ALFA TETFAMGINM PARTVLFTNA RKFDGKDFRW ISSGEYIQMS GRAGRRGMDD RGIVILMVDE KMSPTIGKQL LKGSAD PLN SAFHLTYNMV LNLLRVEEIN PEYMLEKSFY QFQHYRAIPG VVEKVKNSEE QYNKIVIPNE ESVVIYYKIR QQLAKLG KE IEEYIHKPKY CLPFLQPGRL VKVKNEGDDF GWGVVVNFSK KSNVKPNSGE LDPLYVVEVL LRCSKESLKN SATEAAKP A KPDEKGEMQV VPVLVHLLSA ISSVRLYIPK DLRPVDNRQS VLKSIQEVQK RFPDGIPLLD PIDDMGIQDQ GLKKVIQKV EAFEHRMYSH PLHNDPNLET VYTLCEKKAQ IAIDIKSAKR ELKKARTVLQ MDELKCRKRV LRRLGFATSS DVIEMKGRVA CEISSADEL LLTEMMFNGL FNDLSAEQAT ALLSCFVFQE NSSEMPKLTE QLAGPLRQMQ ECAKRIAKVS AEAKLEIDEE T YLSSFKPH LMDVVYTWAT GATFAHICKM TDVFEGSIIR CMRRLEELLR QMCQAAKAIG NTELENKFAE GITKIKRDIV FA ASLYL

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Macromolecule #2: ZCCHC8

MacromoleculeName: ZCCHC8 / type: protein_or_peptide / ID: 2
Details: The first three amino acid residues (SGD) in the sample sequence are cloning artefact.
Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: SGDMAAEVYF GDLELFEPFD HPEESIPKPV HTRFKDDDGD EEDENGVGDA ELRERLRQCE ETIEQLRAEN QELKRKLNIL TRPSGILVN DTKLDGPILQ ILFMNNAISK QYHQEIEEFV SNLVKRFEEQ QKNDVEKTSF NLLPQPSSIV LEEDHKVEES C AIKNNKEA ...String:
SGDMAAEVYF GDLELFEPFD HPEESIPKPV HTRFKDDDGD EEDENGVGDA ELRERLRQCE ETIEQLRAEN QELKRKLNIL TRPSGILVN DTKLDGPILQ ILFMNNAISK QYHQEIEEFV SNLVKRFEEQ QKNDVEKTSF NLLPQPSSIV LEEDHKVEES C AIKNNKEA FSVVGSVLYF TNFCLDKLGQ PLLNENPQLS EGWEIPKYHQ VFSHIVSLEG QEIQVKAKRP KPHCFNCGSE EH QMKDCPM PRNAARISEK RKEYMDACGE ANNQNFQQRY HAEEVEERFG RFKPGVISEE LQDALGVTDK SLPPFIYRMR QLG YPPGWL KEAELENSGL ALYDGKDGTD GETEVGEIQQ NKSVTYDLSK LVNYPGFNIS TPRGIPDEWR IFGSIPMQAC QQKD VFANY LTSNFQAPGV KSGGAVDEDA LTLEELEEQQ RRIWAALEQA ESVNSDSDVP VDTPLTGNSV ASSPCPNELD LPVPE GKTS EKQTLDEPEV PEIFTKKSEA GHASSPDSEV TSLCQKEKAE LAPVNTEGAL LDNGSVVPNC DISNGGSQKL FPADTS PST ATKIHSPIPD MSKFATGITP FEFENMAEST GMYLRIRSLL KNSPRNQQKN KKASE

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Macromolecule #3: RBM7

MacromoleculeName: RBM7 / type: protein_or_peptide / ID: 3
Details: The first three amino acid residues (SGD) in the sample sequence are cloning artefact.
Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString:
SGDEADRTLF VGNLETKVTE ELLFELFHQA GPVIKVKIPK DKDGKPKQFA FVNFKHEVSV PYAMNLLNGI KLYGRPIKIQ FRS

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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
GridModel: UltrAuFoil R1.2/1.3 / Material: GOLD / Mesh: 300 / Support film - Material: GOLD / Pretreatment - Type: GLOW DISCHARGE
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 295 K / Instrument: FEI VITROBOT MARK IV / Details: 30 s wait time, blot for 2.5 s before plunging.

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: SUPER-RESOLUTION / Average electron dose: 67.6 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: OTHER / Details: Ab initio model from RELION
Final reconstructionResolution.type: BY AUTHOR / Resolution: 8.36 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION / Number images used: 52777
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD
FSC plot (resolution estimation)

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