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- EMDB-30639: cryo-EM structure of the DEAH-box helicase Prp2 -

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Basic information

Entry
Database: EMDB / ID: EMD-30639
Titlecryo-EM structure of the DEAH-box helicase Prp2
Map dataEM map of DEAH-box helicase Prp2
Sample
  • Complex: DEAH-box ATPase/helicase Prp2
    • Protein or peptide: PRP2 isoform 1
Keywordsspliceosome / RNA splicing / Bact complex / DEAH-box ATPase/helicase / Prp2 / Spp2 / SPLICING
Function / homology
Function and homology information


snoRNA splicing / generation of catalytic spliceosome for first transesterification step / ATP-dependent activity, acting on RNA / U2-type catalytic step 1 spliceosome / catalytic step 2 spliceosome / helicase activity / nucleic acid binding / RNA helicase activity / RNA helicase / ATP hydrolysis activity ...snoRNA splicing / generation of catalytic spliceosome for first transesterification step / ATP-dependent activity, acting on RNA / U2-type catalytic step 1 spliceosome / catalytic step 2 spliceosome / helicase activity / nucleic acid binding / RNA helicase activity / RNA helicase / ATP hydrolysis activity / RNA binding / ATP binding
Similarity search - Function
: / Helicase associated domain (HA2), ratchet-like / DEAD-box helicase, OB fold / Oligonucleotide/oligosaccharide-binding (OB)-fold / Helicase-associated domain / Helicase associated domain (HA2), winged-helix / Helicase associated domain (HA2) Add an annotation / DNA/RNA helicase, ATP-dependent, DEAH-box type, conserved site / DEAH-box subfamily ATP-dependent helicases signature. / DEAD/DEAH box helicase ...: / Helicase associated domain (HA2), ratchet-like / DEAD-box helicase, OB fold / Oligonucleotide/oligosaccharide-binding (OB)-fold / Helicase-associated domain / Helicase associated domain (HA2), winged-helix / Helicase associated domain (HA2) Add an annotation / DNA/RNA helicase, ATP-dependent, DEAH-box type, conserved site / DEAH-box subfamily ATP-dependent helicases signature. / DEAD/DEAH box helicase / DEAD/DEAH box helicase domain / Helicase conserved C-terminal domain / helicase superfamily c-terminal domain / Superfamilies 1 and 2 helicase C-terminal domain profile. / Superfamilies 1 and 2 helicase ATP-binding type-1 domain profile. / DEAD-like helicases superfamily / Helicase, C-terminal / Helicase superfamily 1/2, ATP-binding domain / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
PRP2 isoform 1 / Pre-mRNA-splicing factor ATP-dependent RNA helicase-like protein PRP2
Similarity search - Component
Biological speciesSaccharomyces cerevisiae (brewer's yeast)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.9 Å
AuthorsBai R / Wan R
Funding support China, 1 items
OrganizationGrant numberCountry
National Natural Science Foundation of China (NSFC)31930059 China
CitationJournal: Science / Year: 2021
Title: Mechanism of spliceosome remodeling by the ATPase/helicase Prp2 and its coactivator Spp2.
Authors: Rui Bai / Ruixue Wan / Chuangye Yan / Qi Jia / Jianlin Lei / Yigong Shi /
Abstract: Spliceosome remodeling, executed by conserved adenosine triphosphatase (ATPase)/helicases including Prp2, enables precursor messenger RNA (pre-mRNA) splicing. However, the structural basis for the ...Spliceosome remodeling, executed by conserved adenosine triphosphatase (ATPase)/helicases including Prp2, enables precursor messenger RNA (pre-mRNA) splicing. However, the structural basis for the function of the ATPase/helicases remains poorly understood. Here, we report atomic structures of Prp2 in isolation, Prp2 complexed with its coactivator Spp2, and Prp2-loaded activated spliceosome and the results of structure-guided biochemical analysis. Prp2 weakly associates with the spliceosome and cannot function without Spp2, which stably associates with Prp2 and anchors on the spliceosome, thus tethering Prp2 to the activated spliceosome and allowing Prp2 to function. Pre-mRNA is loaded into a featured channel between the N and C halves of Prp2, where Leu from the N half and Arg from the C half prevent backward sliding of pre-mRNA toward its 5'-end. Adenosine 5'-triphosphate binding and hydrolysis trigger interdomain movement in Prp2, which drives unidirectional stepwise translocation of pre-mRNA toward its 3'-end. These conserved mechanisms explain the coupling of spliceosome remodeling to pre-mRNA splicing.
History
DepositionOct 26, 2020-
Header (metadata) releaseJan 6, 2021-
Map releaseJan 6, 2021-
UpdateMar 27, 2024-
Current statusMar 27, 2024Processing site: PDBj / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.015
  • Imaged by UCSF Chimera
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  • Surface view colored by radius
  • Surface level: 0.015
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-7dcq
  • Surface level: 0.012
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_30639.png

Downloads & links

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Map

FileDownload / File: emd_30639.map.gz / Format: CCP4 / Size: 103 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationEM map of DEAH-box helicase Prp2
Voxel sizeX=Y=Z: 0.6625 Å
Density
Contour LevelBy AUTHOR: 0.015 / Movie #1: 0.015
Minimum - Maximum-0.038691223 - 0.06761063
Average (Standard dev.)0.000058015947 (±0.0017596649)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions300300300
Spacing300300300
CellA=B=C: 198.75 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z0.66250.66250.6625
M x/y/z300300300
origin x/y/z0.0000.0000.000
length x/y/z198.750198.750198.750
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS300300300
D min/max/mean-0.0390.0680.000

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Supplemental data

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Sample components

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Entire : DEAH-box ATPase/helicase Prp2

EntireName: DEAH-box ATPase/helicase Prp2
Components
  • Complex: DEAH-box ATPase/helicase Prp2
    • Protein or peptide: PRP2 isoform 1

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Supramolecule #1: DEAH-box ATPase/helicase Prp2

SupramoleculeName: DEAH-box ATPase/helicase Prp2 / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Saccharomyces cerevisiae (brewer's yeast)
Molecular weightTheoretical: 101 kDa/nm

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Macromolecule #1: PRP2 isoform 1

MacromoleculeName: PRP2 isoform 1 / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Saccharomyces cerevisiae (brewer's yeast)
Molecular weightTheoretical: 99.947492 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: MSSITSETGK RRVKRTYEVT RQNDNAVRIE PSSLGEEEDK EAKDKNSALQ LKRSRYDPNK VFSNTNQGPE KNNLKGEQLG SQKKSSKYD EKITSNNELT TKKGLLGDSE NETKYASSNS KFNVEVTHKI KNAKEIDKIN RQRMWEEQQL RNAMAGQSDH P DDITLEGS ...String:
MSSITSETGK RRVKRTYEVT RQNDNAVRIE PSSLGEEEDK EAKDKNSALQ LKRSRYDPNK VFSNTNQGPE KNNLKGEQLG SQKKSSKYD EKITSNNELT TKKGLLGDSE NETKYASSNS KFNVEVTHKI KNAKEIDKIN RQRMWEEQQL RNAMAGQSDH P DDITLEGS DKYDYVFDTD AMIDYTNEED DLLPEEKLQY EARLAQALET EEKRILTIQE ARKLLPVHQY KDELLQEIKK NQ VLIIMGE TGSGKTTQLP QYLVEDGFTD QGKLQIAITQ PRRVAATSVA ARVADEMNVV LGKEVGYQIR FEDKTTPNKT VLK YMTDGM LLREFLTDSK LSKYSCIMID EAHERTLATD ILIGLLKDIL PQRPTLKLLI SSATMNAKKF SEFFDNCPIF NVPG RRYPV DIHYTLQPEA NYIHAAITTI FQIHTTQSLP GDILVFLTGQ EEIERTKTKL EEIMSKLGSR TKQMIITPIY ANLPQ EQQL KIFQPTPENC RKVVLATNIA ETSLTIDGIR YVIDPGFVKE NSYVPSTGMT QLLTVPCSRA SVDQRAGRAG RVGPGK CFR IFTKWSYLHE LELMPKPEIT RTNLSNTVLL LLSLGVTDLI KFPLMDKPSI PTLRKSLENL YILGALNSKG TITRLGK MM CEFPCEPEFA KVLYTAATHE QCQGVLEECL TIVSMLHETP SLFIGQKRDA AASVLSEVES DHILYLEIFN QWRNSKFS R SWCQDHKIQF KTMLRVRNIR NQLFRCSEKV GLVEKNDQAR MKIGNIAGYI NARITRCFIS GFPMNIVQLG PTGYQTMGR SSGGLNVSVH PTSILFVNHK EKAQRPSKYV LYQQLMLTSK EFIRDCLVIP KEEWLIDMVP QIFKDLIDDK TNRGRR

UniProtKB: PRP2 isoform 1

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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: 7.9
GridModel: Quantifoil R1.2/1.3 / Material: GOLD / Support film - Material: GRAPHENE / Support film - topology: CONTINUOUS / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 10 sec.
VitrificationCryogen name: ETHANE / Chamber humidity: 100 %

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Electron microscopy

MicroscopeFEI TITAN
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy
Specialist opticsEnergy filter - Slit width: 20 eV
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average electron dose: 50.0 e/Å2

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Image processing

Startup modelType of model: EMDB MAP
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 2.9 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 174455

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