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- PDB-2g4b: Structure of U2AF65 variant with polyuridine tract -

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Basic information

Entry
Database: PDB / ID: 2g4b
TitleStructure of U2AF65 variant with polyuridine tract
Components
  • 5'-R(P*UP*UP*UP*UP*UP*UP*U)-3'
  • Splicing factor U2AF 65 kDa subunit
KeywordsRNA Binding Protein/RNA / protein-RNA complex / RNA splicing factor / RNA recognition motif / RNA Binding Protein-RNA COMPLEX
Function / homology
Function and homology information


U2AF complex / poly-pyrimidine tract binding / pre-mRNA 3'-splice site binding / mRNA 3'-end processing / C2H2 zinc finger domain binding / commitment complex / Transport of Mature mRNA derived from an Intron-Containing Transcript / RNA Polymerase II Transcription Termination / U2-type prespliceosome / molecular function inhibitor activity ...U2AF complex / poly-pyrimidine tract binding / pre-mRNA 3'-splice site binding / mRNA 3'-end processing / C2H2 zinc finger domain binding / commitment complex / Transport of Mature mRNA derived from an Intron-Containing Transcript / RNA Polymerase II Transcription Termination / U2-type prespliceosome / molecular function inhibitor activity / spliceosomal complex assembly / Protein hydroxylation / negative regulation of mRNA splicing, via spliceosome / negative regulation of protein ubiquitination / mRNA Splicing - Major Pathway / positive regulation of RNA splicing / spliceosomal complex / mRNA processing / mRNA splicing, via spliceosome / nuclear speck / enzyme binding / RNA binding / nucleoplasm / nucleus
Similarity search - Function
U2 snRNP auxilliary factor, large subunit, splicing factor / RRM (RNA recognition motif) domain / RNA recognition motif / RNA recognition motif / Eukaryotic RNA Recognition Motif (RRM) profile. / RNA recognition motif domain / RNA-binding domain superfamily / Nucleotide-binding alpha-beta plait domain superfamily / Alpha-Beta Plaits / 2-Layer Sandwich / Alpha Beta
Similarity search - Domain/homology
1,4-DIETHYLENE DIOXIDE / RNA / Splicing factor U2AF 65 kDa subunit
Similarity search - Component
Biological speciesHomo sapiens (human)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 2.5 Å
AuthorsSickmier, E.A. / Kielkopf, C.L.
Citation
Journal: Mol.Cell / Year: 2006
Title: Structural basis of polypyrimidine tract recognition by the essential splicing factor U2AF65.
Authors: Sickmier, E.A. / Frato, K.E. / Paranawithana, S. / Shen, H. / Green, M.R. / Kielkopf, C.L.
#1: Journal: Acta Crystallogr.,Sect.F / Year: 2006
Title: Crystallization and preliminary X-ray analysis of a U2AF65 variant in complex with a polypyrimidine-tract analogue by use of protein engineering.
Authors: Sickmier, E.A. / Frato, K.E. / Kielkopf, C.L.
History
DepositionFeb 21, 2006Deposition site: RCSB / Processing site: RCSB
Revision 1.0Jul 25, 2006Provider: repository / Type: Initial release
Revision 1.1May 1, 2008Group: Version format compliance
Revision 1.2Jul 13, 2011Group: Version format compliance
Revision 1.3Aug 16, 2017Group: Refinement description / Source and taxonomy / Category: entity_src_gen / software
Revision 1.4Jan 31, 2018Group: Database references / Category: citation_author / Item: _citation_author.name
Revision 1.5Aug 30, 2023Group: Data collection / Database references ...Data collection / Database references / Derived calculations / Refinement description
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / database_2 / pdbx_initial_refinement_model / struct_site
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession ..._database_2.pdbx_DOI / _database_2.pdbx_database_accession / _struct_site.pdbx_auth_asym_id / _struct_site.pdbx_auth_comp_id / _struct_site.pdbx_auth_seq_id

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
B: 5'-R(P*UP*UP*UP*UP*UP*UP*U)-3'
A: Splicing factor U2AF 65 kDa subunit
hetero molecules


Theoretical massNumber of molelcules
Total (without water)21,1083
Polymers21,0202
Non-polymers881
Water54030
1
B: 5'-R(P*UP*UP*UP*UP*UP*UP*U)-3'
A: Splicing factor U2AF 65 kDa subunit
hetero molecules

B: 5'-R(P*UP*UP*UP*UP*UP*UP*U)-3'
A: Splicing factor U2AF 65 kDa subunit
hetero molecules


Theoretical massNumber of molelcules
Total (without water)42,2166
Polymers42,0404
Non-polymers1762
Water724
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
crystal symmetry operation5_565y,-x+y+1,z+1/61
Unit cell
Length a, b, c (Å)129.388, 129.388, 62.645
Angle α, β, γ (deg.)90.00, 90.00, 120.00
Int Tables number179
Space group name H-MP6522
DetailsThe second part of the biological assembly is generated by the following six-fold axis: MATRIX 0.5001393 0.8659449 0.0003453 -0.8659446 0.5001394 -0.0007568 -0.0008280 0.0000795 0.9999996 TRANSLATION 129.3170471 -0.0240173 -10.3325310

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Components

#1: RNA chain 5'-R(P*UP*UP*UP*UP*UP*UP*U)-3'


Mass: 2098.203 Da / Num. of mol.: 1 / Source method: obtained synthetically / Details: polypyrimidine tract
#2: Protein Splicing factor U2AF 65 kDa subunit / U2 auxiliary factor 65 kDa subunit / U2 snRNP auxiliary factor large subunit / hU2AF65


Mass: 18921.586 Da / Num. of mol.: 1 / Fragment: RNA binding domain / Mutation: deletion of residues 238-258
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: U2AF65 / Plasmid: pGEX-6p / Species (production host): Escherichia coli / Production host: Escherichia coli BL21 (bacteria) / Strain (production host): BL21 / References: UniProt: P26368
#3: Chemical ChemComp-DIO / 1,4-DIETHYLENE DIOXIDE


Mass: 88.105 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C4H8O2
#4: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 30 / Source method: isolated from a natural source / Formula: H2O

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Experimental details

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Experiment

ExperimentMethod: X-RAY DIFFRACTION / Number of used crystals: 1

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Sample preparation

CrystalDensity Matthews: 3.6 Å3/Da / Density % sol: 65.83 %
Crystal growTemperature: 277 K / Method: vapor diffusion, hanging drop / pH: 6.5
Details: .6 M ammonium sulfate, 100 mM 2-(N-morpholino) ethanesulfonic acid, sodium salt (MES) pH 6.5, 10% dioxane, 200 mM non-detergent sulfobetaine (NDSB) 195 , VAPOR DIFFUSION, HANGING DROP, temperature 277K
Components of the solutions
IDNameCrystal-IDSol-ID
1ammonium sulfate11
22-(N-morpholino) ethanesulfonic acid11
3MES11
4dioxane11
5NDSB11
6H2O11
7ammonium sulfate12
82-(N-morpholino) ethanesulfonic acid12
9MES12
10dioxane12
11NDSB12

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Data collection

DiffractionMean temperature: 100 K
Diffraction sourceSource: SYNCHROTRON / Site: NSLS / Beamline: X8C / Wavelength: 1 Å
DetectorType: ADSC QUANTUM 4 / Detector: CCD / Date: Apr 6, 2005
RadiationMonochromator: parabolic collimating mirror placed upstream of the monochromator
Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 1 Å / Relative weight: 1
ReflectionResolution: 2.5→20 Å / Num. all: 11182 / Num. obs: 11182 / % possible obs: 99.7 % / Observed criterion σ(F): -1 / Observed criterion σ(I): -1
Reflection shellResolution: 2.5→2.59 Å / % possible all: 99.7

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Processing

Software
NameClassification
HKL-2000data collection
SCALEPACKdata scaling
CNSrefinement
HKL-2000data reduction
CNSphasing
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: PDB Entry: 2FZR

2fzr
PDB Unreleased entry


Resolution: 2.5→20 Å / σ(F): 0 / Stereochemistry target values: Engh & Huber
RfactorNum. reflectionSelection details
Rfree0.276 996 random
Rwork0.261 --
all0.306 11137 -
obs0.262 9684 -
Refinement stepCycle: LAST / Resolution: 2.5→20 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms1342 141 6 30 1519
Refine LS restraints
Refine-IDTypeDev ideal
X-RAY DIFFRACTIONc_bond_d0.007
X-RAY DIFFRACTIONc_angle_deg1.51032

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