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Orientation of RNA polymerase II within the human VP16-Mediator-pol II-TFIIF assembly

by single particle reconstruction, at 36 A resolution

Movie

Orientation:

#1: Depositted structure unit, Image by Jmol

#2: Superimposing with simplified surface model of EM map, EMDB-5343, Image by Jmol

#3: Superimposing with EM 3D map: EMDB-5343, Image by UCSF CHIMERA

Entry
Summary
Database / IDPORTEIN DATA BANK (PDB) / 3j0k
TitleOrientation of RNA polymerase II within the human VP16-Mediator-pol II-TFIIF assembly
Descriptor(DNA-directed RNA polymerase II ...) x 7
(DNA-directed RNA polymerases I/II/III ...) x 5
KeywordsTRANSFERASE/TRANSCRIPTION, TRANSFERASE-TRANSCRIPTION complex
AuthorsBernecky, C., Grob, P., Ebmeier, C.C., Nogales, E., Taatjes, D.J.
DateDeposition: 2011-10-04, Release: 2011-10-19
PDBj Mine pagesSummary, Structural Details, Experimental Details, Functional Details
Other databasesRCSB-PDB, PDBe, CATH, CE, FSSP, SCOP, VAST
Sequence detailsTHIS ENTRY WAS MODELED WITH HOMOLOGOUS PROTEIN SEQUENCES FROM SACCHAROMYCES CEREVISIAE.
Structure Visualization
MoviesMovie Page

#1: Depositted structure unit, Image by Jmol

#2: Superimposing with simplified surface model of EM map, EMDB-5343, Image by Jmol

#3: Superimposing with EM 3D map: EMDB-5343, Image by UCSF CHIMERA

Structure viewersYorodumi, jV4, Jmol, Biological unit (Images, jV)
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EMDB-5343

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Article
Citation - primary
ArticlePLoS Biol., Vol. 9, Issue 3, Page e1000603, Year 2011
TitleMolecular architecture of the human Mediator-RNA polymerase II-TFIIF assembly.
AuthorsCarrie Bernecky, Patricia Grob, Christopher C Ebmeier, Eva Nogales, Dylan J Taatjes
Department of Chemistry and Biochemistry, University of Colorado, Boulder, Colorado, United States of America.
KeywordsCryoelectron Microscopy, Humans, Mass Spectrometry, Mediator Complex (chemistry), Models, Molecular, Multiprotein Complexes (chemistry), Protein Conformation, Protein Subunits (chemistry), RNA Polymerase II (chemistry, 2.7.7.-), Transcription Factors, TFII (chemistry), transcription factor TFIIE
LinksDOI: 10.1371/journal.pbio.1000603, PubMed: 21468301, PMC: PMC3066130
Components
ID 1 : DNA-directed RNA polymerase II largest subunit
Image
DescriptionDNA-directed RNA polymerase II largest subunit
Typepolypeptide(L)
Formula weight163180.906 Da
Number of molecules1
ID1
Ec2.7.7.6
SourceMethod: Isolated from a natural source
Common name: human
Cell line: HeLa
NCBI taxonomy: ID:9606
Organelle: nucleus
Organism scientific: Homo sapiens
LinksSequence view
ID 2 : DNA-directed RNA polymerase II 140 kDa polypeptide
Image
DescriptionDNA-directed RNA polymerase II 140 kDa polypeptide
Typepolypeptide(L)
Formula weight138938.062 Da
Number of molecules1
ID2
Ec2.7.7.6
SourceMethod: Isolated from a natural source
Common name: human
Cell line: HeLa
NCBI taxonomy: ID:9606
Organelle: nucleus
Organism scientific: Homo sapiens
LinksSequence view
ID 3 : DNA-directed RNA polymerase II 45 kDa polypeptide
Image
DescriptionDNA-directed RNA polymerase II 45 kDa polypeptide
Typepolypeptide(L)
Formula weight30140.277 Da
Number of molecules1
ID3
Ec2.7.7.6
SourceMethod: Isolated from a natural source
Common name: human
Cell line: HeLa
NCBI taxonomy: ID:9606
Organelle: nucleus
Organism scientific: Homo sapiens
LinksSequence view
ID 4 : DNA-directed RNA polymerase II 32 kDa polypeptide
Image
DescriptionDNA-directed RNA polymerase II 32 kDa polypeptide
Typepolypeptide(L)
Formula weight25451.395 Da
Number of molecules1
ID4
Ec2.7.7.6
SourceMethod: Isolated from a natural source
Common name: human
Cell line: HeLa
NCBI taxonomy: ID:9606
Organelle: nucleus
Organism scientific: Homo sapiens
LinksSequence view
ID 5 : DNA-directed RNA polymerases I, II, and III 27 kDa polypeptide
Image
DescriptionDNA-directed RNA polymerases I, II, and III 27 kDa polypeptide
Typepolypeptide(L)
Formula weight25117.262 Da
Number of molecules1
ID5
Ec2.7.7.6
SourceMethod: Isolated from a natural source
Common name: human
Cell line: HeLa
NCBI taxonomy: ID:9606
Organelle: nucleus
Organism scientific: Homo sapiens
LinksSequence view
ID 6 : DNA-directed RNA polymerases I, II, and III 23 kDa polypeptide
Image
DescriptionDNA-directed RNA polymerases I, II, and III 23 kDa polypeptide
Typepolypeptide(L)
Formula weight9675.320 Da
Number of molecules1
ID6
Ec2.7.7.6
SourceMethod: Isolated from a natural source
Common name: human
Cell line: HeLa
NCBI taxonomy: ID:9606
Organelle: nucleus
Organism scientific: Homo sapiens
LinksSequence view
ID 7 : DNA-directed RNA polymerase II 19 kDa polypeptide
Image
DescriptionDNA-directed RNA polymerase II 19 kDa polypeptide
Typepolypeptide(L)
Formula weight19081.186 Da
Number of molecules1
ID7
Ec2.7.7.6
SourceMethod: Isolated from a natural source
Common name: human
Cell line: HeLa
NCBI taxonomy: ID:9606
Organelle: nucleus
Organism scientific: Homo sapiens
LinksSequence view
ID 8 : DNA-directed RNA polymerases I, II, and III 14.5 kDa polypeptide
Image
DescriptionDNA-directed RNA polymerases I, II, and III 14.5 kDa polypeptide
Typepolypeptide(L)
Formula weight16525.500 Da
Number of molecules1
ID8
Ec2.7.7.6
SourceMethod: Isolated from a natural source
Common name: human
Cell line: HeLa
NCBI taxonomy: ID:9606
Organelle: nucleus
Organism scientific: Homo sapiens
LinksSequence view
ID 9 : DNA-directed RNA polymerase II subunit 9
Image
DescriptionDNA-directed RNA polymerase II subunit 9
Typepolypeptide(L)
Formula weight14308.247 Da
Number of molecules1
ID9
Ec2.7.7.6
SourceMethod: Isolated from a natural source
Common name: human
Cell line: HeLa
NCBI taxonomy: ID:9606
Organelle: nucleus
Organism scientific: Homo sapiens
LinksSequence view
ID 10 : DNA-directed RNA polymerases I/II/III subunit 10
Image
DescriptionDNA-directed RNA polymerases I/II/III subunit 10
Typepolypeptide(L)
Formula weight8290.787 Da
Number of molecules1
ID10
Ec2.7.7.6
SourceMethod: Isolated from a natural source
Common name: human
Cell line: HeLa
NCBI taxonomy: ID:9606
Organelle: nucleus
Organism scientific: Homo sapiens
LinksSequence view
ID 11 : DNA-directed RNA polymerase II 13.6 kDa polypeptide
Image
DescriptionDNA-directed RNA polymerase II 13.6 kDa polypeptide
Typepolypeptide(L)
Formula weight13633.628 Da
Number of molecules1
ID11
Ec2.7.7.6
SourceMethod: Isolated from a natural source
Common name: human
Cell line: HeLa
NCBI taxonomy: ID:9606
Organelle: nucleus
Organism scientific: Homo sapiens
LinksSequence view
ID 12 : DNA-directed RNA polymerases I, II, and III 7.7 kDa polypeptide
Image
DescriptionDNA-directed RNA polymerases I, II, and III 7.7 kDa polypeptide
Typepolypeptide(L)
Formula weight5252.292 Da
Number of molecules1
ID12
Ec2.7.7.6
SourceMethod: Isolated from a natural source
Common name: human
Cell line: HeLa
NCBI taxonomy: ID:9606
Organelle: nucleus
Organism scientific: Homo sapiens
LinksSequence view
ID 13 : MAGNESIUM ION
Image
DescriptionMAGNESIUM ION
Typenon-polymer
Formula weight24.305 Da
Number of molecules1
ID13
SourceMethod: Obtained synthetically
ID 14 : ZINC ION
Image
DescriptionZINC ION
Typenon-polymer
Formula weight65.380 Da
Number of molecules9
ID14
SourceMethod: Obtained synthetically
Sample
Assembly
Aggregation statePARTICLE
Compositionone Mediator complex binds one pol II-TFIIF
Theoretical Mass1.9
Num components3
Entity assembly
Assembly-ID1
Ebi cellHeLa
Ebi organelleNucleus
Ebi organism commonhuman
Ebi organism scientificHomo sapiens
Mutant flagNO
Namecore Mediator
Oligomeric details26 subunit complex
SynonymMediator
TypePROTEIN
Entity assembly
Assembly-ID1
Ebi cellHeLa
Ebi organelleNucleus
Ebi organism commonHuman
Ebi organism scientificHomo sapiens
Mutant flagNO
NameRNA polymerase II
Oligomeric details12-subunit complex
Synonympol II
TypePROTEIN
Entity assembly
Assembly-ID1
Ebi expression systemEscherichia coli
Ebi organism commonHuman
Ebi organism scientificHomo sapiens
Mutant flagYES
NameTFIIF
Oligomeric detailsDimer
SynonymTFIIF
TypePROTEIN
Experiment
Reconstruction methodSINGLE PARTICLE
Specimen typeVITREOUS ICE (CRYO EM)
Sample preparation
Details20 mM HEPES, 0.10 mM EDTA, 150 mM KCl, 0.02% NP-40, 35% glycerol
pH7.9
Sample support
Detailsthin carbon-coated holey carbon 400 mesh copper grid
Vitrification
Cryogen nameETHANE
Methodblot for 2 seconds, dry for 3 seconds before plunging
Temp90 Kelvin
Experiment
MethodELECTRON MICROSCOPY
Electron Microscopy
Imaging
MicroscopeModel: FEI TECNAI F20
Electron gun
Electron sourceFIELD EMISSION GUN
Accelerating voltage200 kV
Electron dose15 e/A**2
Illumination modeFLOOD BEAM
Lens
ModeBRIGHT FIELD
MagnificationNominal: 29000 X
CsNominal: 2.0 mm
Detector distance0.0
Nominal defocusMax: 4500 nm, Min: 1000 nm
Specimen holder
Specimen holderModel: GATAN LIQUID NITROGEN, Type: side entry
Tilt angleMin: 0.0 degrees, Max: 0.0 degrees
Detector
TypeKodak SO-163 film
Image scans
Number digital images106
Od range1.0
Quant bit size16
Sampling size12.9
Scanner modelOTHER
Processing
2D projection selection
Software nameSitus
Single particle entity
Symmetry typeASYMMETRIC
3D reconstruction
CTF correction methodeach micrograph
DetailsThe particles were selected interactively.
Methodmulti-reference projection matching
Number of particles3146
Resolution36 A
Resolution methodFSC at 0.5 cut-off
SoftwareSpider
3D fitting
Detailsthe TFIIS chain S was removed before fitting
Refinement Protocolrigid body
Refinement SpaceREAL
Software nameSitus
Target criteriacontour-based Laplacian correlation
3D fitting list
3D Fitting ID1
PDB entry ID1Y1V
Refine hist
Cycle idLAST
Refine idELECTRON MICROSCOPY
Total atoms31137
Ligand atoms10
Protein atoms31127
Download
PDB format
Allpdb3j0k.ent.gz
pdb3j0k.ent (uncompressed file)
Header onlypdb3j0k.ent.gz
mmCIF format
mmCIF3j0k.cif.gz
XML format
All3j0k.xml.gz
No-atom3j0k-noatom.xml.gz
Ext-atom3j0k-extatom.xml.gz
Movie files
movie #1
.mp4 (H.264/MPEG-4 AVC format), 2.8 MB
.webm (WebM/VP8 format), 3.7 MB
movie #2
.mp4 (H.264/MPEG-4 AVC format), 3.4 MB
.webm (WebM/VP8 format), 4.4 MB
movie #3
.mp4 (H.264/MPEG-4 AVC format), 3.4 MB
.webm (WebM/VP8 format), 4.4 MB