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- PDB-5xjc: Cryo-EM structure of the human spliceosome just prior to exon lig... -

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Entry
Database: PDB / ID: 5xjc
TitleCryo-EM structure of the human spliceosome just prior to exon ligation at 3.6 angstrom
DescriptorPre-mRNA-processing-splicing factor 8
116 kDa U5 small nuclear ribonucleoprotein component
Crooked neck-like protein 1
Cell division cycle 5-like protein
Protein BUD31 homolog
and more 18 items
KeywordsSPLICING / RNA splicing / human spliceosome / C* complex / atomic structure / step 2 factors / EJC
Specimen sourceHomo sapiens / human
Human adenovirus 2 / virus
MethodElectron microscopy (3.6 Å resolution / Particle / Single particle)
AuthorsZhang, X. / Yan, C. / Hang, J. / Finci, I.L. / Lei, J. / Shi, Y.
CitationCell, 2017, 169, 918-929.e14

Cell, 2017, 169, 918-929.e14 StrPapers
An Atomic Structure of the Human Spliceosome.
Xiaofeng Zhang / Chuangye Yan / Jing Hang / Lorenzo I Finci / Jianlin Lei / Yigong Shi

Validation Report
SummaryFull reportAbout validation report
DateDeposition: Apr 30, 2017 / Release: Jul 5, 2017

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Assembly

Deposited unit
A: Pre-mRNA-processing-splicing factor 8
B: U5 snRNA
C: 116 kDa U5 small nuclear ribonucleoprotein component
D: U5 small nuclear ribonucleoprotein 200 kDa helicase
E: U5 small nuclear ribonucleoprotein 40 kDa protein
F: U6 snRNA
G: pre-mRNA
H: Homo sapiens small nuclear RNA (U2) gene
I: Pre-mRNA-splicing factor SYF1
J: Crooked neck-like protein 1
K: Pre-mRNA-splicing factor SPF27
L: Cell division cycle 5-like protein
M: Pre-mRNA-splicing factor SYF2
N: Protein BUD31 homolog
O: Pre-mRNA-splicing factor RBM22
P: Spliceosome-associated protein CWC15 homolog
Q: Intron-binding protein aquarius
R: SNW domain-containing protein 1
S: Peptidyl-prolyl cis-trans isomerase-like 1
T: Pleiotropic regulator 1
U: Serine/arginine repetitive matrix protein 2
V: Pre-mRNA-splicing factor CWC22 homolog
W: Pre-mRNA-processing factor 17
X: PRKR-interacting protein 1
Y: ATP-dependent RNA helicase DHX8
Z: Pre-mRNA-splicing factor SLU7
a: Small nuclear ribonucleoprotein Sm D3
b: Small nuclear ribonucleoprotein-associated proteins B and B'
c: Small nuclear ribonucleoprotein Sm D1
d: Small nuclear ribonucleoprotein Sm D2
f: Small nuclear ribonucleoprotein F
e: Small nuclear ribonucleoprotein E
g: Small nuclear ribonucleoprotein G
h: Small nuclear ribonucleoprotein Sm D3
i: Small nuclear ribonucleoprotein-associated proteins B and B'
j: Small nuclear ribonucleoprotein Sm D1
k: Small nuclear ribonucleoprotein Sm D2
m: Small nuclear ribonucleoprotein F
l: Small nuclear ribonucleoprotein E
n: Small nuclear ribonucleoprotein G
o: U2 small nuclear ribonucleoprotein A'
p: U2 small nuclear ribonucleoprotein B''
q: Pre-mRNA-processing factor 19
r: Pre-mRNA-processing factor 19
s: Pre-mRNA-processing factor 19
t: Pre-mRNA-processing factor 19
u: Eukaryotic initiation factor 4A-III
v: Protein mago nashi homolog 2
w: RNA-binding protein 8A
x: Protein CASC3
hetero molecules


Theoretical massNumber of molelcules
Total (without water)2,962,08074
Polyers2,958,30350
Non-polymers3,77724
Water0
#1


  • idetical with deposited unit
  • defined by author
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1

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Components

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Polypeptide(L) , 18 types, 19 molecules ACJLNPQRST...

#1: Polypeptide(L)Pre-mRNA-processing-splicing factor 8 / 220 kDa U5 snRNP-specific protein / PRP8 homolog / Splicing factor Prp8 / p220


Mass: 273974.250 Da / Num. of mol.: 1 / Source: (natural) Homo sapiens / References: UniProt: Q6P2Q9
#3: Polypeptide(L)116 kDa U5 small nuclear ribonucleoprotein component / Elongation factor Tu GTP-binding domain-containing protein 2 / SNU114 homolog / hSNU114 / U5 snRNP-specific protein / 116 kDa / U5-116 kDa


Mass: 109560.625 Da / Num. of mol.: 1 / Source: (natural) Homo sapiens / References: UniProt: Q15029
#10: Polypeptide(L)Crooked neck-like protein 1 / Crooked neck homolog / hCrn


Mass: 100610.008 Da / Num. of mol.: 1 / Source: (natural) Homo sapiens / References: UniProt: Q9BZJ0
#12: Polypeptide(L)Cell division cycle 5-like protein / Cdc5-like protein / Pombe cdc5-related protein


Mass: 92406.883 Da / Num. of mol.: 1 / Source: (natural) Homo sapiens / References: UniProt: Q99459
#14: Polypeptide(L)Protein BUD31 homolog / Protein EDG-2 / Protein G10 homolog


Mass: 17032.850 Da / Num. of mol.: 1 / Source: (natural) Homo sapiens / References: UniProt: P41223
#16: Polypeptide(L)Spliceosome-associated protein CWC15 homolog


Mass: 26674.447 Da / Num. of mol.: 1 / Source: (natural) Homo sapiens / References: UniProt: Q9P013
#17: Polypeptide(L)Intron-binding protein aquarius / Intron-binding protein of 160 kDa / IBP160


Mass: 171502.453 Da / Num. of mol.: 1 / Source: (natural) Homo sapiens / References: UniProt: O60306
#18: Polypeptide(L)SNW domain-containing protein 1 / Nuclear protein SkiP / Nuclear receptor coactivator NCoA-62 / Ski-interacting protein


Mass: 61770.648 Da / Num. of mol.: 1 / Source: (natural) Homo sapiens / References: UniProt: Q13573
#19: Polypeptide(L)Peptidyl-prolyl cis-trans isomerase-like 1 / PPIase / Rotamase PPIL1


Mass: 18257.805 Da / Num. of mol.: 1 / Source: (natural) Homo sapiens / References: UniProt: Q9Y3C6, EC: 5.2.1.8
#20: Polypeptide(L)Pleiotropic regulator 1


Mass: 57280.758 Da / Num. of mol.: 1 / Source: (natural) Homo sapiens / References: UniProt: O43660
#21: Polypeptide(L)Serine/arginine repetitive matrix protein 2 / 300 kDa nuclear matrix antigen / Serine/arginine-rich splicing factor-related nuclear matrix protein of 300 kDa / Ser/Arg-related nuclear matrix protein of 300 kDa / Splicing coactivator subunit SRm300 / Tax-responsive enhancer element-binding protein 803 / TaxREB803


Mass: 300255.312 Da / Num. of mol.: 1 / Source: (natural) Homo sapiens / References: UniProt: Q9UQ35
#24: Polypeptide(L)PRKR-interacting protein 1


Mass: 21040.176 Da / Num. of mol.: 1 / Source: (natural) Homo sapiens / References: UniProt: Q9H875
#25: Polypeptide(L)ATP-dependent RNA helicase DHX8 / DEAH box protein 8 / RNA helicase HRH1


Mass: 139508.328 Da / Num. of mol.: 1 / Source: (natural) Homo sapiens / References: UniProt: Q14562, EC: 3.6.4.13
#28: Polypeptide(L)Small nuclear ribonucleoprotein-associated proteins B and B' / snRNP-B / Sm protein B/B' / SmB/B'


Mass: 23686.004 Da / Num. of mol.: 2 / Source: (natural) Homo sapiens / References: UniProt: P14678
#37: Polypeptide(L)Eukaryotic initiation factor 4A-III / eIF4A-III / ATP-dependent RNA helicase DDX48 / ATP-dependent RNA helicase eIF4A-3 / DEAD box protein 48 / Eukaryotic initiation factor 4A-like NUK-34 / Eukaryotic translation initiation factor 4A isoform 3 / Nuclear matrix protein 265 / hNMP 265


Mass: 46930.961 Da / Num. of mol.: 1 / Source: (natural) Homo sapiens / References: UniProt: P38919, EC: 3.6.4.13
#38: Polypeptide(L)Protein mago nashi homolog 2


Mass: 17301.799 Da / Num. of mol.: 1 / Source: (natural) Homo sapiens / References: UniProt: Q96A72
#39: Polypeptide(L)RNA-binding protein 8A / Binder of OVCA1-1 / BOV-1 / RNA-binding motif protein 8A / RNA-binding protein Y14 / Ribonucleoprotein RBM8A


Mass: 19925.070 Da / Num. of mol.: 1 / Source: (natural) Homo sapiens / References: UniProt: Q9Y5S9
#40: Polypeptide(L)Protein CASC3 / Cancer susceptibility candidate gene 3 protein / Metastatic lymph node gene 51 protein / MLN 51 / Protein barentsz / Btz


Mass: 76381.992 Da / Num. of mol.: 1 / Source: (natural) Homo sapiens / References: UniProt: O15234

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RNA chain , 4 types, 4 molecules BFGH

#2: RNA chainU5 snRNA


Mass: 37254.855 Da / Num. of mol.: 1 / Source: (natural) Homo sapiens
#6: RNA chainU6 snRNA


Mass: 34404.438 Da / Num. of mol.: 1 / Source: (natural) Homo sapiens
#7: RNA chainpre-mRNA


Mass: 87977.836 Da / Num. of mol.: 1 / Source: (synth.) Human adenovirus 2
#8: RNA chainHomo sapiens small nuclear RNA (U2) gene


Mass: 60186.445 Da / Num. of mol.: 1 / Source: (natural) Homo sapiens / References: GenBank: 609501

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U5 small nuclear ribonucleoprotein ... , 2 types, 2 molecules DE

#4: Polypeptide(L)U5 small nuclear ribonucleoprotein 200 kDa helicase / Activating signal cointegrator 1 complex subunit 3-like 1 / BRR2 homolog / U5 snRNP-specific 200 kDa protein / U5-200KD


Mass: 244823.422 Da / Num. of mol.: 1 / Source: (natural) Homo sapiens / References: UniProt: O75643, EC: 3.6.4.13
#5: Polypeptide(L)U5 small nuclear ribonucleoprotein 40 kDa protein / U5-40K / 38 kDa-splicing factor / Prp8-binding protein / hPRP8BP / U5 snRNP-specific 40 kDa protein / WD repeat-containing protein 57


Mass: 39359.492 Da / Num. of mol.: 1 / Source: (natural) Homo sapiens / References: UniProt: Q96DI7

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Pre-mRNA-splicing factor ... , 6 types, 6 molecules IKMOVZ

#9: Polypeptide(L)Pre-mRNA-splicing factor SYF1 / Protein HCNP / XPA-binding protein 2


Mass: 100148.711 Da / Num. of mol.: 1 / Source: (natural) Homo sapiens / References: UniProt: Q9HCS7
#11: Polypeptide(L)Pre-mRNA-splicing factor SPF27 / Breast carcinoma-amplified sequence 2 / DNA amplified in mammary carcinoma 1 protein / Spliceosome-associated protein SPF 27


Mass: 26163.420 Da / Num. of mol.: 1 / Source: (natural) Homo sapiens / References: UniProt: O75934
#13: Polypeptide(L)Pre-mRNA-splicing factor SYF2 / CCNDBP1-interactor / p29


Mass: 28780.518 Da / Num. of mol.: 1 / Source: (natural) Homo sapiens / References: UniProt: O95926
#15: Polypeptide(L)Pre-mRNA-splicing factor RBM22 / RNA-binding motif protein 22 / Zinc finger CCCH domain-containing protein 16


Mass: 46959.555 Da / Num. of mol.: 1 / Source: (natural) Homo sapiens / References: UniProt: Q9NW64
#22: Polypeptide(L)Pre-mRNA-splicing factor CWC22 homolog / Nucampholin homolog / fSAPb


Mass: 105646.578 Da / Num. of mol.: 1 / Source: (natural) Homo sapiens / References: UniProt: Q9HCG8
#26: Polypeptide(L)Pre-mRNA-splicing factor SLU7 / hSlu7


Mass: 68510.844 Da / Num. of mol.: 1 / Source: (natural) Homo sapiens / References: UniProt: O95391

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Pre-mRNA-processing factor ... , 2 types, 5 molecules Wqrst

#23: Polypeptide(L)Pre-mRNA-processing factor 17 / Cell division cycle 40 homolog / EH-binding protein 3 / Ehb3 / PRP17 homolog / hPRP17


Mass: 65612.180 Da / Num. of mol.: 1 / Source: (natural) Homo sapiens / References: UniProt: O60508
#36: Polypeptide(L)
Pre-mRNA-processing factor 19 / Nuclear matrix protein 200 / PRP19/PSO4 homolog / hPso4 / RING-type E3 ubiquitin transferase PRP19 / Senescence evasion factor


Mass: 55245.547 Da / Num. of mol.: 4 / Source: (natural) Homo sapiens / References: UniProt: Q9UMS4, EC: 2.3.2.27

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Small nuclear ribonucleoprotein ... , 6 types, 12 molecules ahcjdkfmelgn

#27: Polypeptide(L)Small nuclear ribonucleoprotein Sm D3 / Sm-D3 / snRNP core protein D3


Mass: 13940.308 Da / Num. of mol.: 2 / Source: (natural) Homo sapiens / References: UniProt: P62318
#29: Polypeptide(L)Small nuclear ribonucleoprotein Sm D1 / Sm-D1 / Sm-D autoantigen / snRNP core protein D1


Mass: 13310.653 Da / Num. of mol.: 2 / Source: (natural) Homo sapiens / References: UniProt: P62314
#30: Polypeptide(L)Small nuclear ribonucleoprotein Sm D2 / Sm-D2 / snRNP core protein D2


Mass: 13551.928 Da / Num. of mol.: 2 / Source: (natural) Homo sapiens / References: UniProt: P62316
#31: Polypeptide(L)Small nuclear ribonucleoprotein F / snRNP-F / Sm protein F / SmF


Mass: 9734.171 Da / Num. of mol.: 2 / Source: (natural) Homo sapiens / References: UniProt: P62306
#32: Polypeptide(L)Small nuclear ribonucleoprotein E / snRNP-E / Sm protein E / SmE


Mass: 10817.601 Da / Num. of mol.: 2 / Source: (natural) Homo sapiens / References: UniProt: P62304
#33: Polypeptide(L)Small nuclear ribonucleoprotein G / snRNP-G / Sm protein G / SmG


Mass: 8508.084 Da / Num. of mol.: 2 / Source: (natural) Homo sapiens / References: UniProt: P62308

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U2 small nuclear ribonucleoprotein ... , 2 types, 2 molecules op

#34: Polypeptide(L)U2 small nuclear ribonucleoprotein A' / U2 snRNP A'


Mass: 28456.584 Da / Num. of mol.: 1 / Source: (natural) Homo sapiens / References: UniProt: P09661
#35: Polypeptide(L)U2 small nuclear ribonucleoprotein B'' / U2 snRNP B''


Mass: 25524.367 Da / Num. of mol.: 1 / Source: (natural) Homo sapiens / References: UniProt: P08579

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Non-polymers , 6 types, 24 molecules

#41: ChemicalChemComp-I6P / INOSITOL 1,2,3,4,5,6-HEXAKISPHOSPHATE / MYO-INOSITOL HEXAKISPHOSPHATE


Mass: 660.035 Da / Num. of mol.: 1 / Formula: C6H18O24P6
#42: ChemicalChemComp-GTP / GUANOSINE-5'-TRIPHOSPHATE / GTP *YM


Mass: 523.180 Da / Num. of mol.: 1 / Formula: C10H16N5O14P3
#43: Chemical
ChemComp-MG / MAGNESIUM ION


Mass: 24.305 Da / Num. of mol.: 11 / Formula: Mg
#44: ChemicalChemComp-ADP / ADENOSINE-5'-DIPHOSPHATE / ADP *YM


Mass: 427.201 Da / Num. of mol.: 2 / Formula: C10H15N5O10P2
#45: Chemical
ChemComp-ZN / ZINC ION


Mass: 65.409 Da / Num. of mol.: 7 / Formula: Zn
#46: ChemicalChemComp-ATP / ADENOSINE-5'-TRIPHOSPHATE / ATP (energy-carrying molecule) *YM


Mass: 507.181 Da / Num. of mol.: 2 / Formula: C10H16N5O13P3

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Details

Sequence detailsThe UNK reisue between G-1 and G1 is added intentionally as per authors request to fill in the cleavage by RNA splicing.

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / Reconstruction method: SINGLE PARTICLE

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Sample preparation

ComponentName: human C* spliceosome / Type: COMPLEX
Entity ID: 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43
Source: NATURAL
Source (natural)Organism: Homo sapiens
Buffer solutionpH: 7.5
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
MicroscopyMicroscope model: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Cs: 2.7 mm / Alignment procedure: BASIC
Specimen holderCryogen: NITROGEN / Specimen holder model: GATAN LIQUID NITROGEN
Image recordingAverage exposure time: 0.25 sec. / Electron dose: 50 e/Å2 / Detector mode: SUPER-RESOLUTION / Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Number of grids imaged: 5 / Number of real images: 7308
EM imaging opticsEnergyfilter name: Gatan Quantum 967 LS / Energyfilter upper: 10 eV / Energyfilter lower: 0 eV / Chr aberration corrector: -10
Image scansMovie frames/image: 32 / Used frames/image: 2-32

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Processing

EM software
IDNameVersionCategoryImage processing ID
10RELION2.0INITIAL EULER ASSIGNMENT1
11RELION2.0FINAL EULER ASSIGNMENT1
13RELION2.0RECONSTRUCTION1
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 3.6 Å / Resolution method: FSC 0.143 CUT-OFF / Number of particles: 100085 / Symmetry type: POINT
Least-squares processHighest resolution: 3.6 Å

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