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- EMDB-51225: 5'-lobe of the substrate-bound U11 snRNP -

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Basic information

Entry
Database: EMDB / ID: EMD-51225
Title5'-lobe of the substrate-bound U11 snRNP
Map datamain map
Sample
  • Complex: 5'-lobe of the substrate-bound U11 snRNP
    • Protein or peptide: U11/U12 small nuclear ribonucleoprotein 25 kDa protein
    • Protein or peptide: U11/U12 small nuclear ribonucleoprotein 35 kDa protein
    • RNA: P120-5'SS
    • RNA: U11 snRNA
Keywordsminor spliceosome / U11 snRNP / RNA-protein complex / SPLICING
Function / homology
Function and homology information


snRNA binding / U12-type spliceosomal complex / mRNA Splicing - Minor Pathway / intercellular bridge / RNA splicing / mRNA splicing, via spliceosome / mRNA binding / nucleolus / nucleoplasm / nucleus / cytosol
Similarity search - Function
snRNP35, RNA recognition motif / U11/U12 small nuclear ribonucleoprotein 25kDa protein / SNRNP25, ubiquitin-like domain / Ubiquitin-like domain / : / RNA recognition motif / RNA recognition motif / Eukaryotic RNA Recognition Motif (RRM) profile. / RNA recognition motif domain / RNA-binding domain superfamily ...snRNP35, RNA recognition motif / U11/U12 small nuclear ribonucleoprotein 25kDa protein / SNRNP25, ubiquitin-like domain / Ubiquitin-like domain / : / RNA recognition motif / RNA recognition motif / Eukaryotic RNA Recognition Motif (RRM) profile. / RNA recognition motif domain / RNA-binding domain superfamily / Ubiquitin domain profile. / Ubiquitin-like domain / Ubiquitin-like domain superfamily / Nucleotide-binding alpha-beta plait domain superfamily
Similarity search - Domain/homology
U11/U12 small nuclear ribonucleoprotein 35 kDa protein / U11/U12 small nuclear ribonucleoprotein 25 kDa protein
Similarity search - Component
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.4 Å
AuthorsZhao J / Galej WP
Funding supportEuropean Union, 1 items
OrganizationGrant numberCountry
European Research Council (ERC)950278European Union
CitationJournal: Mol Cell / Year: 2025
Title: Structural basis of 5' splice site recognition by the minor spliceosome.
Authors: Jiangfeng Zhao / Daniel Peter / Irina Brandina / Xiangyang Liu / Wojciech P Galej /
Abstract: The minor spliceosome catalyzes excision of U12-dependent introns from precursors of eukaryotic messenger RNAs (pre-mRNAs). This process is critical for many cellular functions, but the underlying ...The minor spliceosome catalyzes excision of U12-dependent introns from precursors of eukaryotic messenger RNAs (pre-mRNAs). This process is critical for many cellular functions, but the underlying molecular mechanisms remain elusive. Here, we report a cryoelectron microscopy (cryo-EM) reconstruction of the 13-subunit human U11 small nuclear ribonucleoprotein particle (snRNP) complex in apo and substrate-bound forms, revealing the architecture of the U11 small nuclear RNA (snRNA), five minor spliceosome-specific factors, and the mechanism of the U12-type 5' splice site (5'SS) recognition. SNRNP25 and SNRNP35 specifically recognize U11 snRNA, while PDCD7 bridges SNRNP25 and SNRNP48, located at the distal ends of the particle. SNRNP48 and ZMAT5 are positioned near the 5' end of U11 snRNA and stabilize binding of the incoming 5'SS. Recognition of the U12-type 5'SS is achieved through base-pairing to the 5' end of the U11 snRNA and unexpected, non-canonical base-triple interactions with the U11 snRNA stem-loop 3. Our structures provide mechanistic insights into U12-dependent intron recognition and the evolution of the splicing machinery.
History
DepositionJul 31, 2024-
Header (metadata) releaseFeb 5, 2025-
Map releaseFeb 5, 2025-
UpdateFeb 19, 2025-
Current statusFeb 19, 2025Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_51225.png

Downloads & links

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Map

FileDownload / File: emd_51225.map.gz / Format: CCP4 / Size: 325 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotationmain map
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.73 Å/pix.
x 440 pix.
= 321.2 Å		0.73 Å/pix.
x 440 pix.
= 321.2 Å		0.73 Å/pix.
x 440 pix.
= 321.2 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.73 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.03
Minimum - Maximum-0.079917565 - 0.14640306
Average (Standard dev.)0.00004318046 (±0.0025444657)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions440440440
Spacing440440440
CellA=B=C: 321.2 Å
α=β=γ: 90.0 °

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Supplemental data

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Additional map: sharp map

Fileemd_51225_additional_1.map
Annotationsharp map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: half A

Fileemd_51225_half_map_1.map
Annotationhalf A
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: half B

Fileemd_51225_half_map_2.map
Annotationhalf B
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : 5'-lobe of the substrate-bound U11 snRNP

EntireName: 5'-lobe of the substrate-bound U11 snRNP
Components
  • Complex: 5'-lobe of the substrate-bound U11 snRNP
    • Protein or peptide: U11/U12 small nuclear ribonucleoprotein 25 kDa protein
    • Protein or peptide: U11/U12 small nuclear ribonucleoprotein 35 kDa protein
    • RNA: P120-5'SS
    • RNA: U11 snRNA

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Supramolecule #1: 5'-lobe of the substrate-bound U11 snRNP

SupramoleculeName: 5'-lobe of the substrate-bound U11 snRNP / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 100 KDa

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Macromolecule #1: U11/U12 small nuclear ribonucleoprotein 25 kDa protein

MacromoleculeName: U11/U12 small nuclear ribonucleoprotein 25 kDa protein
type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 15.290729 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString:
MDVFQEGLAM VVQDPLLCDL PIQVTLEEVN SQIALEYGQA MTVRVCKMDG EVMPVVVVQS ATVLDLKKAI QRYVQLKQER EGGIQHISW SYVWRTYHLT SAGEKLTEDR KKLRDYGIRN RDEVSFIKKL RQK

UniProtKB: U11/U12 small nuclear ribonucleoprotein 25 kDa protein

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Macromolecule #2: U11/U12 small nuclear ribonucleoprotein 35 kDa protein

MacromoleculeName: U11/U12 small nuclear ribonucleoprotein 35 kDa protein
type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 29.514471 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: MNDWMPIAKE YDPLKAGSID GTDEDPHDRA VWRAMLARYV PNKGVIGDPL LTLFVARLNL QTKEDKLKEV FSRYGDIRRL RLVRDLVTG FSKGYAFIEY KEERAVIKAY RDADGLVIDQ HEIFVDYELE RTLKGWIPRR LGGGLGGKKE SGQLRFGGRD R PFRKPINL ...String:
MNDWMPIAKE YDPLKAGSID GTDEDPHDRA VWRAMLARYV PNKGVIGDPL LTLFVARLNL QTKEDKLKEV FSRYGDIRRL RLVRDLVTG FSKGYAFIEY KEERAVIKAY RDADGLVIDQ HEIFVDYELE RTLKGWIPRR LGGGLGGKKE SGQLRFGGRD R PFRKPINL PVVKNDLYRE GKRERRERSR SRERHWDSRT RDRDHDRGRE KRWQEREPTR VWPDNDWERE RDFRDDRIKG RE KKERGK

UniProtKB: U11/U12 small nuclear ribonucleoprotein 35 kDa protein

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Macromolecule #3: P120-5'SS

MacromoleculeName: P120-5'SS / type: rna / ID: 3 / Number of copies: 1
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 3.366978 KDa
SequenceString:
AUAUCCUUUU U

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Macromolecule #4: U11 snRNA

MacromoleculeName: U11 snRNA / type: rna / ID: 4 / Number of copies: 1
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 43.505629 KDa
SequenceString:
AAAAAGGGCU UCUGUCGUGA GUGGCACACG UAGGGCAACU CGAUUGCUCU GCGUGCGGAA UCGACAUCAA GAGAUUUCGG AAGCAUAAU UUUUUGGUAU UUGGGCAGCU GGUGAUCGUU GGUCCCGGCG CCCUUU

GENBANK: GENBANK: NR_004407.1

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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

Concentration0.2 mg/mL
BufferpH: 7.9
Component:
ConcentrationFormulaName
200.0 mMKClpotassium chloride
2.0 mMMgCl2magnesium chloride

Details: 20 mM HEPES-KOH, pH 7.9 200 mM KCl 2 mM MgCl2
GridModel: UltrAuFoil R2/2 / Pretreatment - Type: PLASMA CLEANING / Pretreatment - Time: 90 sec. / Pretreatment - Atmosphere: OTHER
VitrificationCryogen name: ETHANE / Chamber humidity: 95 % / Chamber temperature: 277.15 K / Instrument: FEI VITROBOT MARK IV
DetailsThis sample was monodisperse

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Specialist opticsEnergy filter - Name: TFS Selectris X / Energy filter - Slit width: 10 eV
DetailsPreliminary grid screening was performed on Glacios.
Image recordingFilm or detector model: FEI FALCON IV (4k x 4k) / Digitization - Dimensions - Width: 4096 pixel / Digitization - Dimensions - Height: 4096 pixel / Number grids imaged: 1 / Number real images: 20000 / Average electron dose: 41.73 e/Å2 / Details: Images were colleted in EER format
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 50.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 1.5 µm / Nominal defocus min: 0.7000000000000001 µm / Nominal magnification: 165000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 4494715 / Details: Topaz picking is performed with user-trained model
Startup modelType of model: PDB ENTRY
PDB model - PDB ID:

8r7n

Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.4 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. v4.3.1) / Number images used: 52290
Initial angle assignmentType: RANDOM ASSIGNMENT / Software - Name: cryoSPARC (ver. v4.3.1)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. v4.3.1)
Final 3D classificationNumber classes: 5 / Avg.num./class: 22588 / Software - Name: cryoSPARC (ver. v4.3.1)
Details: The heterogeneous refinement was performed to separate particles.

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Atomic model buiding 1

Initial model
PDB IDChain

0-f1

chain_id: B, residue_range: 1-132, source_name: AlphaFold, initial_model_type: in silico model

0-f1

chain_id: C, residue_range: 10-164, source_name: AlphaFold, initial_model_type: in silico model
Detailsrefinement is done in Phenix
RefinementSpace: REAL / Protocol: RIGID BODY FIT / Overall B value: 73.3 / Target criteria: Cross-correction coefficient
Output model

PDB-9gbz:
5'-lobe of the substrate-bound U11 snRNP

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