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- PDB-7luv: Cryo-EM structure of the yeast THO-Sub2 complex -

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Basic information

Entry
Database: PDB / ID: 7luv
TitleCryo-EM structure of the yeast THO-Sub2 complex
Components
  • (THO complex subunit ...) x 4
  • ATP-dependent RNA helicase SUB2
  • Tex1
KeywordsRNA BINDING PROTEIN / nuclear mRNA export / DEAD-box ATPase / mRNP remodeling
Function / homology
Function and homology information


nucleoplasmic THO complex / THO complex part of transcription export complex / transcription export complex / Cdc73/Paf1 complex / positive regulation of transcription elongation from RNA polymerase I promoter / positive regulation of transcription by RNA polymerase I / mRNA 3'-end processing / chromatin silencing at telomere / stress granule assembly / molecular adaptor activity ...nucleoplasmic THO complex / THO complex part of transcription export complex / transcription export complex / Cdc73/Paf1 complex / positive regulation of transcription elongation from RNA polymerase I promoter / positive regulation of transcription by RNA polymerase I / mRNA 3'-end processing / chromatin silencing at telomere / stress granule assembly / molecular adaptor activity / transcription elongation from RNA polymerase II promoter / mRNA export from nucleus / spliceosomal complex / telomere maintenance / mRNA splicing, via spliceosome / transcription-coupled nucleotide-excision repair / mRNA processing / chromosome, telomeric region / nucleic acid binding / RNA helicase / DNA recombination / RNA helicase activity / mRNA binding / RNA binding / ATP binding / nucleus / cytoplasm
DEAD/DEAH box helicase / THO complex, subunitTHOC2, N-terminal / Helicase, C-terminal / THO complex subunit 7/Mft1 / DEAD/DEAH box helicase domain / Helicase superfamily 1/2, ATP-binding domain / RNA helicase, DEAD-box type, Q motif / THO complex, subunitTHOC2, C-terminal / THO complex subunit Thp2 / THO complex, subunit THOC1 ...DEAD/DEAH box helicase / THO complex, subunitTHOC2, N-terminal / Helicase, C-terminal / THO complex subunit 7/Mft1 / DEAD/DEAH box helicase domain / Helicase superfamily 1/2, ATP-binding domain / RNA helicase, DEAD-box type, Q motif / THO complex, subunitTHOC2, C-terminal / THO complex subunit Thp2 / THO complex, subunit THOC1 / P-loop containing nucleoside triphosphate hydrolase / THO complex subunit 2, N-terminal domain / THO complex subunit 2 / Tho complex subunit THP2
THO complex subunit THP2 / THO complex subunit HPR1 / THO complex subunit MFT1 / THO complex subunit 2 / ATP-dependent RNA helicase SUB2
Biological speciesSaccharomyces cerevisiae (baker's yeast)
Saccharomyces bayanus (yeast)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.7 Å
AuthorsXie, Y. / Ren, Y.
Funding support United States, 1items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM133743 United States
CitationJournal: Elife / Year: 2021
Title: Cryo-EM structure of the yeast TREX complex and coordination with the SR-like protein Gbp2.
Authors: Yihu Xie / Bradley P Clarke / Yong Joon Kim / Austin L Ivey / Pate S Hill / Yi Shi / Yi Ren /
Abstract: The evolutionarily conserved TRanscript-EXport (TREX) complex plays central roles during mRNP (messenger ribonucleoprotein) maturation and export from the nucleus to the cytoplasm. In yeast, TREX is ...The evolutionarily conserved TRanscript-EXport (TREX) complex plays central roles during mRNP (messenger ribonucleoprotein) maturation and export from the nucleus to the cytoplasm. In yeast, TREX is composed of the THO sub-complex (Tho2, Hpr1, Tex1, Mft1, and Thp2), the DEAD box ATPase Sub2, and Yra1. Here we present a 3.7 Å cryo-EM structure of the yeast THO•Sub2 complex. The structure reveals the intimate assembly of THO revolving around its largest subunit Tho2. THO stabilizes a semi-open conformation of the Sub2 ATPase via interactions with Tho2. We show that THO interacts with the serine-arginine (SR)-like protein Gbp2 through both the RS domain and RRM domains of Gbp2. Cross-linking mass spectrometry analysis supports the extensive interactions between THO and Gbp2, further revealing that RRM domains of Gbp2 are in close proximity to the C-terminal domain of Tho2. We propose that THO serves as a landing pad to configure Gbp2 to facilitate its loading onto mRNP.
Validation Report
SummaryFull reportAbout validation report
History
DepositionFeb 23, 2021Deposition site: RCSB / Processing site: RCSB
Revision 1.0Apr 14, 2021Provider: repository / Type: Initial release

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Structure visualization

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Assembly

Deposited unit
A: THO complex subunit HPR1
B: THO complex subunit THP2
C: THO complex subunit 2
D: THO complex subunit MFT1
E: Tex1
M: ATP-dependent RNA helicase SUB2


Theoretical massNumber of molelcules
Total (without water)361,4406
Polymers361,4406
Non-polymers00
Water0
1


TypeNameSymmetry operationNumber
identity operation1_5551
Buried area35670 Å2
ΔGint-233 kcal/mol
Surface area118080 Å2

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Components

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THO complex subunit ... , 4 types, 4 molecules ABCD

#1: Protein THO complex subunit HPR1 / Hyperrecombination protein 1


Mass: 71126.195 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharomyces cerevisiae (baker's yeast)
Gene: HPR1, YDR138W, YD9302.14 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: P17629
#2: Protein THO complex subunit THP2


Mass: 30340.264 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharomyces cerevisiae (baker's yeast)
Gene: THP2, YHR167W / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: O13539
#3: Protein THO complex subunit 2 / Low dye-binding protein 5 / THO complex subunit RLR1 / Zinc-regulated gene 13 protein


Mass: 138881.453 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharomyces cerevisiae (baker's yeast)
Gene: THO2, LDB5, RLR1, ZRG13, YNL139C, N1209, N1835 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: P53552
#4: Protein THO complex subunit MFT1 / Mitochondrial fusion target protein 1


Mass: 29467.480 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharomyces cerevisiae (baker's yeast)
Gene: MFT1, YML062C / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: P33441

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Protein , 2 types, 2 molecules EM

#5: Protein Tex1


Mass: 41248.668 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharomyces bayanus (yeast) / Production host: Trichoplusia ni (cabbage looper)
#6: Protein ATP-dependent RNA helicase SUB2 / Suppressor of BRR1 protein 2


Mass: 50375.859 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharomyces cerevisiae (baker's yeast)
Gene: SUB2, YDL084W / Production host: Escherichia coli (E. coli) / References: UniProt: Q07478, RNA helicase

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Details

Sequence detailsCertain segments of chains C and E were unidentifiable, and so they were modeled as poly-UNK. The ...Certain segments of chains C and E were unidentifiable, and so they were modeled as poly-UNK. The full sequence of chain C is: GAMGSMAEQTLLSKLNALSQKVIPPASPSQASILTEEVIRNWPERSKTLCSDFTALESNDEKEDWLRTLFIELFDFINKN DENSPLKLSDVASFTNELVNHERQVSQASIVGKMFIAVSSTVPNINDLTTISLCKLIPSLHEELFKFSWISSKLLNKEQT TLLRHLLKKSKYELKKYNLLVENSVGYGQLVALLILAYYDPDNFSKVSAYLKEIYHIMGKYSLDSIRTLDVILNVSSQFI TEGYKFFIALLRKSDSWPSSHVANNSNYSSLNEGGNMIAANIISFNLSQYNEEVDKENYERYMDMCCILLKNGFVNFYSI WDNVKPEMEFLQEYIQNLETELEEESTKGVENPLAMAAALSTENETDEDNALVVNDDVNMKDKISEETNADIESKGKQKT QQDILLFGKIKLLERLLIHGCVIPVIHVLKQYPKVLYVSESLSRYLGRVFEYLLNPLYTSMTSSGESKDMATALMITRID NGILAHKPRLIHKYKTHEPFESLELNSSYVFYYSEWNSNLTPFASVNDLFENSHIYLSIIGPYLGRIPTLLSKISRIGVA DIQKNHGSESLHVTIDKWIDYVRKFIFPATSLLQNNPIATSEVYELMKFFPFEKRYFIYNEMMTKLSQDILPLKVSFNKA EREAKSILKALSIDTIAKESRRFAKLISTNPLASLVPAVKQIENYDKVSELVVYTTKYFNDFAYDVLQFVLLLRLTYNRP AVQFDGVNQAMWVQRLSIFIAGLAKNCPNMDISNIITYILKTLHNGNIIAVSILKELIITVGGIRDLNEVNMKQLLMLNS GSPLKQYARHLIYDFRDDNSVISSRLTSFFTDQSAISEIILLLYTLNLKANTQNSHYKILSTRCDEMNTLLWSFIELIKH CLKGKAFEENVLPFVELNNRFHLSTPWTFHIWRDYLDNQLNSNENFSIDELIEGAEFSDVDLTKISKDLFTTFWRLSLYD IHFDKSLYDERKNALSGENTGHMSNRKKHLIQNQIKDILVTGISHQRAFKKTSEFISEKS NVWNKDCGEDQIKIFLQNCVVPRVLFSPSDALFSSFFIFMAFRTENLMSILNTCITSNILKTLLFCCTSSEAGNLGLFFTDVLKKLEKMRLNGD FNDQASRKLYEWHSVITEQVIDLLSEKNYMSIRNGIEFMKHVTSVFPVVKAHIQLVYTTLEENLINEEREDIKLPSSALI GHLKARLKDALELDEFCTLTEEEAEQKRIREMELEEIKNYETACQNEQKQVALRKQLELNKSQRLQND The full sequence of chain E is: GAMGSMPSSETIDIVNQKTITSEVAASVTSKYLQSTLSKNNISDIEDERFIHVSSRSHSRFTSTPVTPNEVLSLKFHASG SSMAYSRMDGSLTVWFIKDASFDKSVKVYIPDCCGSDKLATDLSWNPTSLNQMAVVSNSSEISLLLINEITLTASKLRTL SLGSKTKVNSCLYDPLGNWLLAATKSEKIYLFNVKEDHRLVHSLNVNDISPNTNDVVYSIAWNNNGSHIFVGFKSGHLVI LKLRDEMLEVSTKIKAHTSSITGIKIDPWGRYFVTGSSDGNCYIWSLKSLCCEKIIQDLDSAVVALDVCHLGKILGVCTE DEMVYFYDLNEAKLLESKSLANHKSDLVLKFYPDKSWYILSGKNDTLSNHFVKNEKNLIT YWKDM

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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

Component
IDNameTypeEntity IDParent-IDSource
1THO-Sub2COMPLEX#1-#60MULTIPLE SOURCES
2Sub2COMPLEX#61RECOMBINANT
3THOCOMPLEX#1-#41RECOMBINANT
4Tex1COMPLEX#51RECOMBINANT
Source (natural)
IDEntity assembly-IDOrganismNcbi tax-ID
12Saccharomyces cerevisiae (baker's yeast)4932
23Saccharomyces cerevisiae (baker's yeast)4932
34Saccharomyces bayanus (yeast)4931
Source (recombinant)
IDEntity assembly-IDOrganismNcbi tax-ID
32E. coli (E. coli)562
43Trichoplusia ni (cabbage looper)7111
54Trichoplusia ni (cabbage looper)7111
Buffer solutionpH: 7
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy
Image recordingElectron dose: 50 e/Å2 / Film or detector model: FEI FALCON III (4k x 4k)

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Processing

SoftwareName: PHENIX / Version: 1.11.1_2575: / Classification: refinement
EM softwareName: PHENIX / Version: 1.11.1 / Category: model refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 3.7 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 30066 / Symmetry type: POINT
Atomic model buildingB value: 114.5 / Protocol: AB INITIO MODEL / Space: REAL
Refine LS restraints
Refinement-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00719116
ELECTRON MICROSCOPYf_angle_d0.74525844
ELECTRON MICROSCOPYf_dihedral_angle_d8.7211570
ELECTRON MICROSCOPYf_chiral_restr0.0462969
ELECTRON MICROSCOPYf_plane_restr0.0043259

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