EMDB-12974: Structure of active transcription elongation complex Pol II-DSIF ...

Basic information

• Entry: Database: EMDB / ID: EMD-12974
• Title: Structure of active transcription elongation complex Pol II-DSIF (SPT5-KOW5) alone, Map 11, Global map
• Map data: Map 11, main map for Pol II-DSIF (SPT5-KOW5) alone model

Sample

• Complex: Transcription elongation complex of RNA polymerase II with elongation factors DSIF (SPT5-KOW5)
  • Protein or peptide: x 13 types
  • RNA: x 1 types
  • DNA: x 2 types
• Ligand: x 2 types

• Keywords: Pol II / Super elongation complex / ELL2 / EAF1 / TRANSCRIPTION

Function / homology

Function:

negative regulation of DNA-templated transcription, elongation / : / DSIF complex / regulation of transcription elongation by RNA polymerase II / B-WICH complex positively regulates rRNA expression / RNA Polymerase I Transcription Initiation / RNA Polymerase I Promoter Escape / RNA Polymerase I Transcription Termination / RNA Polymerase III Transcription Initiation From Type 1 Promoter / RNA Polymerase III Transcription Initiation From Type 2 Promoter / RNA Polymerase III Transcription Initiation From Type 3 Promoter / Formation of RNA Pol II elongation complex / Formation of the Early Elongation Complex / Transcriptional regulation by small RNAs / RNA Polymerase II Pre-transcription Events / TP53 Regulates Transcription of DNA Repair Genes / FGFR2 alternative splicing / RNA polymerase II transcribes snRNA genes / mRNA Capping / mRNA Splicing - Minor Pathway / Processing of Capped Intron-Containing Pre-mRNA / RNA Polymerase II Promoter Escape / RNA Polymerase II Transcription Pre-Initiation And Promoter Opening / RNA Polymerase II Transcription Initiation / RNA Polymerase II Transcription Elongation / RNA Polymerase II Transcription Initiation And Promoter Clearance / RNA Pol II CTD phosphorylation and interaction with CE / Estrogen-dependent gene expression / Formation of TC-NER Pre-Incision Complex / Dual incision in TC-NER / Gap-filling DNA repair synthesis and ligation in TC-NER / mRNA Splicing - Major Pathway / positive regulation of DNA-templated transcription, elongation / Abortive elongation of HIV-1 transcript in the absence of Tat / transcription elongation-coupled chromatin remodeling / RNA Pol II CTD phosphorylation and interaction with CE during HIV infection / RNA Pol II CTD phosphorylation and interaction with CE / Formation of the Early Elongation Complex / Formation of the HIV-1 Early Elongation Complex / mRNA Capping / organelle membrane / RNA polymerase III activity / maintenance of transcriptional fidelity during transcription elongation by RNA polymerase II / Pausing and recovery of Tat-mediated HIV elongation / Tat-mediated HIV elongation arrest and recovery / positive regulation of macroautophagy / positive regulation of nuclear-transcribed mRNA poly(A) tail shortening / HIV elongation arrest and recovery / Pausing and recovery of HIV elongation / RNA polymerase II transcribes snRNA genes / RNA polymerase II activity / Tat-mediated elongation of the HIV-1 transcript / transcription elongation by RNA polymerase I / Formation of HIV-1 elongation complex containing HIV-1 Tat / transcription-coupled nucleotide-excision repair / tRNA transcription by RNA polymerase III / RNA polymerase I activity / RNA polymerase I complex / RNA polymerase III complex / positive regulation of translational initiation / Formation of HIV elongation complex in the absence of HIV Tat / RNA polymerase II, core complex / RNA Polymerase II Transcription Elongation / Formation of RNA Pol II elongation complex / RNA Polymerase II Pre-transcription Events / translation initiation factor binding / TP53 Regulates Transcription of DNA Repair Genes / transcription initiation at RNA polymerase II promoter / transcription elongation by RNA polymerase II / P-body / ribonucleoside binding / fibrillar center / DNA-directed 5'-3' RNA polymerase activity / DNA-directed RNA polymerase / single-stranded DNA binding / transcription by RNA polymerase II / nucleic acid binding / chromosome, telomeric region / single-stranded RNA binding / protein dimerization activity / nuclear speck / protein heterodimerization activity / RNA-dependent RNA polymerase activity / mRNA binding / nucleotide binding / DNA-templated transcription / chromatin binding / nucleolus / negative regulation of transcription by RNA polymerase II / enzyme binding / positive regulation of transcription by RNA polymerase II / DNA binding / RNA binding / zinc ion binding / nucleoplasm / nucleus / metal ion binding / cytosol

Domain/homology:

Spt5, KOW domain repeat 6 / Transcription elongation factor Spt5, eukaryote / Spt5 transcription elongation factor, N-terminal / Spt5, KOW domain repeat 2 / Spt5, KOW domain repeat 3 / Spt5, KOW domain repeat 5 / Spt5 transcription elongation factor, acidic N-terminal / NGN domain, eukaryotic / Spt5, KOW domain repeat 1 / Spt5, KOW domain repeat 4 / Spt5 C-terminal domain / Spt5 C-terminal nonapeptide repeat binding Spt4 / NGN domain / Transcription elongation factor SPT5 / Early transcription elongation factor of RNA pol II, NGN section / RNA-binding domain, S1 / NusG, N-terminal / In Spt5p, this domain may confer affinity for Spt4p. It possesses a RNP-like fold. / NusG, N-terminal domain superfamily / DNA-directed RNA polymerase II subunit Rpb4-like / RNA polymerase Rpb4/RPC9, core / DNA-directed RNA-polymerase II subunit / Rpb4/RPC9 superfamily / Pol II subunit B9, C-terminal zinc ribbon / RNA polymerase RBP11 / RNA polymerase subunit Rpb4/RPC9 / RNA polymerase Rpb4 / Zinc finger TFIIS-type signature. / HRDC-like superfamily / RNA polymerase Rpb7-like , N-terminal / RNA polymerase Rpb7-like, N-terminal domain superfamily / RNA polymerase subunit Rpb7-like / SHS2 domain found in N terminus of Rpb7p/Rpc25p/MJ0397 / RNA polymerase Rpb2, domain 4 / RNA polymerase Rpb2, domain 4 / RNA polymerase Rpb2, domain 5 / RNA polymerase Rpb2, domain 5 / DNA-directed RNA polymerase, M/15kDa subunit / RNA polymerases M/15 Kd subunit / RNA polymerase subunit 9 / DNA-directed RNA polymerase subunit RPABC5/Rpb10 / RNA polymerases, subunit N, zinc binding site / RNA polymerase subunit RPB10 / RNA polymerases N / 8 kDa subunit / RNA polymerases N / 8 Kd subunits signature. / DNA-directed RNA polymerase M, 15kDa subunit, conserved site / RNA polymerases M / 15 Kd subunits signature. / DNA-directed RNA polymerase subunit/transcription factor S / : / RNA polymerase, Rpb8 / DNA-directed RNA polymerases I, II, and III subunit RPABC4 / RNA polymerase Rpb8 / RNA polymerase subunit 8 / RNA polymerase, Rpb5, N-terminal / RNA polymerase Rpb5, N-terminal domain superfamily / RNA polymerase Rpb5, N-terminal domain / DNA-directed RNA polymerase, subunit RPB6 / DNA directed RNA polymerase, 7 kDa subunit / RNA polymerase archaeal subunit P/eukaryotic subunit RPABC4 / RNA polymerase, subunit H/Rpb5, conserved site / RNA polymerases H / 23 Kd subunits signature. / RNA polymerase subunit CX / DNA-directed RNA polymerase, 30-40kDa subunit, conserved site / DNA-directed RNA polymerase subunit Rpo3/Rpb3/RPAC1 / RNA polymerases D / 30 to 40 Kd subunits signature. / DNA-directed RNA polymerase Rpb11, 13-16kDa subunit, conserved site / DNA-directed RNA polymerase subunit Rpo11 / RNA polymerases L / 13 to 16 Kd subunits signature. / Zinc finger, TFIIS-type / DNA-directed RNA polymerase subunit Rpo5/Rpb5 / Transcription factor S-II (TFIIS) / Zinc finger TFIIS-type profile. / C2C2 Zinc finger / DNA-directed RNA polymerase, RBP11-like dimerisation domain / RNA polymerase Rpb3/Rpb11 dimerisation domain / RNA polymerase, subunit H/Rpb5 C-terminal / RNA polymerase subunit RPABC4/transcription elongation factor Spt4 / RPB5-like RNA polymerase subunit superfamily / RNA polymerase Rpb5, C-terminal domain / Archaeal Rpo6/eukaryotic RPB6 RNA polymerase subunit / DNA-directed RNA polymerase, 14-18kDa subunit, conserved site / RNA polymerases K / 14 to 18 Kd subunits signature. / Ribosomal protein S1-like RNA-binding domain / S1 RNA binding domain / S1 domain / RNA polymerase Rpb6 / RNA polymerase, subunit omega/Rpo6/RPB6 / RNA polymerase Rpb6 / RPB6/omega subunit-like superfamily / RNA polymerase Rpb2, domain 2 superfamily / RNA polymerase, beta subunit, protrusion / RNA polymerase beta subunit / DNA-directed RNA polymerase, insert domain / DNA-directed RNA polymerase, RpoA/D/Rpb3-type / RNA polymerase Rpb3/RpoA insert domain / RNA polymerase Rpb3/Rpb11 dimerisation domain / RNA polymerases D / DNA-directed RNA polymerase, insert domain superfamily / RNA polymerase, RBP11-like subunit / RNA polymerase Rpb2, domain 2

Component:

RNA polymerase II subunit D / DNA-directed RNA polymerase II subunit RPB11-a / DNA-directed RNA polymerases I, II, and III subunit RPABC3 / DNA-directed RNA polymerases I, II, and III subunit RPABC5 / DNA-directed RNA polymerases I, II, and III subunit RPABC2 / DNA-directed RNA polymerase II subunit RPB3 / DNA-directed RNA polymerase subunit beta / DNA-directed RNA polymerase II subunit RPB7 / RNA polymerase II, I and III subunit K / DNA-directed RNA polymerase II subunit E / Transcription elongation factor SPT5 / DNA-directed RNA polymerase II subunit RPB9

• Biological species: Sus scrofa (pig) / Homo sapiens (human) / HIV whole-genome vector AA1305#18 (others)
• Method: single particle reconstruction / cryo EM / Resolution: 3.0 Å
• Authors: Chen Y / Vos SM / Dienemann C / Ninov M / Urlaub H / Cramer P

Funding support

Germany, 4 items

Organization	Grant number	Country
German Research Foundation (DFG)	SFB860	Germany
German Research Foundation (DFG)	EXC 2067/1-390729940	Germany
European Research Council (ERC)	Advanced Grant CHROMATRANS 693023	Germany
German Research Foundation (DFG)	SFB1286	Germany

• Citation: Journal: Mol Cell / Year: 2021; Title: Allosteric transcription stimulation by RNA polymerase II super elongation complex.; Authors: Ying Chen / Seychelle M Vos / Christian Dienemann / Momchil Ninov / Henning Urlaub / Patrick Cramer / ; Abstract: The super elongation complex (SEC) contains the positive transcription elongation factor b (P-TEFb) and the subcomplex ELL2-EAF1, which stimulates RNA polymerase II (RNA Pol II) elongation. Here, we report the cryoelectron microscopy (cryo-EM) structure of ELL2-EAF1 bound to a RNA Pol II elongation complex at 2.8 Å resolution. The ELL2-EAF1 dimerization module directly binds the RNA Pol II lobe domain, explaining how SEC delivers P-TEFb to RNA Pol II. The same site on the lobe also binds the initiation factor TFIIF, consistent with SEC binding only after the transition from transcription initiation to elongation. Structure-guided functional analysis shows that the stimulation of RNA elongation requires the dimerization module and the ELL2 linker that tethers the module to the RNA Pol II protrusion. Our results show that SEC stimulates elongation allosterically and indicate that this stimulation involves stabilization of a closed conformation of the RNA Pol II active center cleft.

History

Deposition	May 18, 2021	-
Header (metadata) release	Jul 14, 2021	-
Map release	Jul 14, 2021	-
Update	Jul 10, 2024	-
Current status	Jul 10, 2024	Processing site: PDBe / Status: Released

Map

• File: Download / File: emd_12974.map.gz / Format: CCP4 / Size: 103 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
• Annotation: Map 11, main map for Pol II-DSIF (SPT5-KOW5) alone model
• Voxel size: X=Y=Z: 1.05 Å

Density

Contour Level	By AUTHOR: 0.018 / Movie #1: 0.018
Minimum - Maximum	-0.08875522 - 0.15978946
Average (Standard dev.)	0.00019237849 (±0.005338669)

• Symmetry: Space group: 1

Details

EMDB XML:

Map geometry	Axis order X Y Z Origin 0 0 0 Dimensions 300 300 300 Spacing 300 300 300
Cell	A=B=C: 315.0 Å α=β=γ: 90.0 °

CCP4 map header:

mode	Image stored as Reals
Å/pix. X/Y/Z	1.05	1.05	1.05
M x/y/z	300	300	300
origin x/y/z	0.000	0.000	0.000
length x/y/z	315.000	315.000	315.000
α/β/γ	90.000	90.000	90.000
start NX/NY/NZ	0	0	0
NX/NY/NZ	300	300	300
MAP C/R/S	1	2	3
start NC/NR/NS	0	0	0
NC/NR/NS	300	300	300
D min/max/mean	-0.089	0.160	0.000

Supplemental data

Mask #1

• File: emd_12974_msk_1.map

Half map: half map 1 for map 11

• File: emd_12974_half_map_1.map
• Annotation: half map 1 for map 11

Half map: half map 2 for map 11

• File: emd_12974_half_map_2.map
• Annotation: half map 2 for map 11

Sample components

Entire : Transcription elongation complex of RNA polymerase II with elonga...

• Entire: Name: Transcription elongation complex of RNA polymerase II with elongation factors DSIF (SPT5-KOW5)

Components

• Complex: Transcription elongation complex of RNA polymerase II with elongation factors DSIF (SPT5-KOW5)
  • Protein or peptide: DNA-directed RNA polymerase II subunit RPB1
  • Protein or peptide: DNA-directed RNA polymerase subunit beta
  • Protein or peptide: DNA-directed RNA polymerase II subunit RPB3
  • Protein or peptide: RNA polymerase II subunit D
  • Protein or peptide: DNA-directed RNA polymerase II subunit E
  • Protein or peptide: DNA-directed RNA polymerase II subunit F
  • Protein or peptide: DNA-directed RNA polymerase II subunit RPB7
  • Protein or peptide: DNA-directed RNA polymerases I, II, and III subunit RPABC3
  • Protein or peptide: DNA-directed RNA polymerase II subunit RPB9
  • Protein or peptide: DNA-directed RNA polymerases I, II, and III subunit RPABC5
  • Protein or peptide: RNA_pol_L_2 domain-containing protein
  • Protein or peptide: RNA polymerase II subunit K
  • Protein or peptide: Transcription elongation factor SPT5
  • RNA: RNA
  • DNA: Non-template DNA
  • DNA: Template DNA
• Ligand: MAGNESIUM ION
• Ligand: ZINC ION

Supramolecule #1: Transcription elongation complex of RNA polymerase II with elonga...

• Supramolecule: Name: Transcription elongation complex of RNA polymerase II with elongation factors DSIF (SPT5-KOW5); type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#16; Details: This is a for map for Pol II-DSIF alone class. In this map, ELL2, EAF1, SPT4 are not visible. For SPT5, only the KOW5 domain is visible.
• Source (natural): Organism: Sus scrofa (pig)

Macromolecule #1: DNA-directed RNA polymerase II subunit RPB1

• Macromolecule: Name: DNA-directed RNA polymerase II subunit RPB1 / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
• Source (natural): Organism: Sus scrofa (pig)
• Molecular weight: Theoretical: 217.450078 KDa

Sequence

String:

MHGGGPPSGD SACPLRTIKR VQFGVLSPDE LKRMSVTEGG IKYPETTEGG RPKLGGLMDP RQGVIERTGR CQTCAGNMTE CPGHFGHIE LAKPVFHVGF LVKTMKVLRC VCFFCSKLLV DSNNPKIKDI LAKSKGQPKK RLTHVYDLCK GKNICEGGEE M DNKFGVEQ PEGDEDLTKE KGHGGCGRYQ PRIRRSGLEL YAEWKHVNED SQEKKILLSP ERVHEIFKRI SDEECFVLGM EP RYARPEW MIVTVLPVPP LSVRPAVVMQ GSARNQDDLT HKLADIVKIN NQLRRNEQNG AAAHVIAEDV KLLQFHVATM VDN ELPGLP RAMQKSGRPL KSLKQRLKGK EGRVRGNLMG KRVDFSARTV ITPDPNLSID QVGVPRSIAA NMTFAEIVTP FNID RLQEL VRRGNSQYPG AKYIIRDNGD RIDLRFHPKP SDLHLQTGYK VERHMCDGDI VIFNRQPTLH KMSMMGHRVR ILPWS TFRL NLSVTTPYNA DFDGDEMNLH LPQSLETRAE IQELAMVPRM IVTPQSNRPV MGIVQDTLTA VRKFTKRDVF LERGEV MNL LMFLSTWDGK VPQPAILKPR PLWTGKQIFS LIIPGHINCI RTHSTHPDDE DSGPYKHISP GDTKVVVENG ELIMGIL CK KSLGTSAGSL VHISYLEMGH DITRLFYSNI QTVINNWLLI EGHTIGIGDS IADSKTYQDI QNTIKKAKQD VIEVIEKA H NNELEPTPGN TLRQTFENQV NRILNDARDK TGSSAQKSLS EYNNFKSMVV SGAKGSKINI SQVIAVVGQQ NVEGKRIPF GFKHRTLPHF IKDDYGPESR GFVENSYLAG LTPTEFFFHA MGGREGLIDT AVKTAETGYI QRRLIKSMES VMVKYDATVR NSINQVVQL RYGEDGLAGE SVEFQNLATL KPSNKAFEKK FRFDYTNERA LRRTLQEDLV KDVLSNAHIQ NELEREFERM R EDREVLRV IFPTGDSKVV LPCNLLRMIW NAQKIFHINP RLPSDLHPIK VVEGVKELSK KLVIVNGDDP LSRQAQENAT LL FNIHLRS TLCSRRMAEE FRLSGEAFDW LLGEIESKFN QAIAHPGEMV GALAAQSLGE PATQMTLNTF HYAGVSAKNV TLG VPRLKE LINISKKPKT PSLTVFLLGQ SARDAERAKD ILCRLEHTTL RKVTANTAIY YDPNPQSTVV AEDQEWVNVY YEMP DFDVA RISPWLLRVE LDRKHMTDRK LTMEQIAEKI NAGFGDDLNC IFNDDNAEKL VLRIRIMNSD ENKMQEEEEV VDKMD DDVF LRCIESNMLT DMTLQGIEQI SKVYMHLPQT DNKKKIIITE DGEFKALQEW ILETDGVSLM RVLSEKDVDP VRTTSN DIV EIFTVLGIEA VRKALERELY HVISFDGSYV NYRHLALLCD TMTCRGHLMA ITRHGVNRQD TGPLMKCSFE ETVDVLM EA AAHGESDPMK GVSENIMLGQ LAPAGTGCFD LLLDAEKCKY GMEIPTNIPG LGAAGPTGMF FGSAPSPMGG ISPAMTPW N QGATPAYGAW SPSVGSGMTP GAAGFSPSAA SDASGFSPGY SPAWSPTPGS PGSPGPSSPY IPSPGGAMSP SYSPTSPAY EPRSPGGYTP QSPSYSPTSP SYSPTSPSYS PTSPNYSPTS PSYSPTSPSY SPTSPSYSPT SPSYSPTSPS YSPTSPSYSP TSPSYSPTS PSYSPTSPSY SPTSPSYSPT SPSYSPTSPS YSPTSPSYSP TSPSYSPTSP SYSPTSPNYS PTSPNYTPTS P SYSPTSPS YSPTSPNYTP TSPNYSPTSP SYSPTSPSYS PTSPSYSPSS PRYTPQSPTY TPSSPSYSPS SPSYSPTSPK YT PTSPSYS PSSPEYTPTS PKYSPTSPKY SPTSPKYSPT SPTYSPTTPK YSPTSPTYSP TSPVYTPTSP KYSPTSPTYS PTS PKYSPT SPTYSPTSPK GSTYSPTSPG YSPTSPTYSL TSPAISPDDS DEEN

Macromolecule #2: DNA-directed RNA polymerase subunit beta

• Macromolecule: Name: DNA-directed RNA polymerase subunit beta / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO / EC number: DNA-directed RNA polymerase
• Source (natural): Organism: Sus scrofa (pig)
• Molecular weight: Theoretical: 142.426125 KDa

Sequence

String:

MCSTNLSQAL AYFRAGANLT AASLWAGFRG SRRIFWERRK RKSLCLALRP GGACWRWLLA VSCVSLRGLG APGSCANMYD ADEDMQYDE DDDEITPDLW QEACWIVISS YFDEKGLVRQ QLDSFDEFIQ MSVQRIVEDA PPIDLQAEAQ HASGEVEEPP R YLLKFEQI YLSKPTHWER DGAPSPMMPN EARLRNLTYS APLYVDITKT VIKEGEEQLQ TQHQKTFIGK IPIMLRSTYC LL NGLTDRD LCELNECPLD PGGYFIINGS EKVLIAQEKM ATNTVYVFAK KDSKYAYTGE CRSCLENSSR PTSTIWVSML ARG GQGAKK SAIGQRIVAT LPYIKQEVPI IIVFRALGFV SDRDILEHII YDFEDPEMME MVKPSLDEAF VIQEQNVALN FIGS RGAKP GVTKEKRIKY AKEVLQKEML PHVGVSDFCE TKKAYFLGYM VHRLLLAALG RRELDDRDHY GNKRLDLAGP LLAFL FRGM FKNLLKEVRI YAQKFIDRGK DFNLELAIKT RIISDGLKYS LATGNWGDQK KAHQARAGVS QVLNRLTFAS TLSHLR RLN SPIGRDGKLA KPRQLHNTLW GMVCPAETPE GHAVGLVKNL ALMAYISVGS QPSPILEFLE EWSMENLEEI SPAAIAD AT KIFVNGCWVG IHKDPEQLMN TLRKLRRQMD IIVSEVSMIR DIREREIRIY TDAGRICRPL LIVEKQKLLL KKRHIDQL K EREYNNYSWQ DLVASGVVEY IDTLEEETVM LAMTPDDLQE KEVAYCSTYT HCEIHPSMIL GVCASIIPFP DHNQSPRNT YQSAMGKQAM GVYITNFHVR MDTLAHVLYY PQKPLVTTRS MEYLRFRELP AGINSIVAIA SYTGYNQEDS VIMNRSAVDR GFFRSVFYR SYKEQESKKG FDQEEVFEKP TRETCQGMRH AIYDKLDDDG LIAPGVRVSG DDVIIGKTVT LPENEDELEG T NRRYTKRD CSTFLRTSET GIVDQVMVTL NQEGYKFCKI RVRSVRIPQI GDKFASRHGQ KGTCGIQYRQ EDMPFTCEGI TP DIIINPH AIPSRMTIGH LIECLQGKVS ANKGEIGDAT PFNDAVNVQK ISNLLSDYGY HLRGNEVLYN GFTGRKITSQ IFI GPTYYQ RLKHMVDDKI HSRARGPIQI LNRQPMEGRS RDGGLRFGEM ERDCQIAHGA AQFLRERLFE ASDPYQVHVC NLCG IMAIA NTRTHTYECR GCRNKTQISL VRMPYACKLL FQELMSMSIA PRMMSV

UniProtKB: DNA-directed RNA polymerase subunit beta

Macromolecule #3: DNA-directed RNA polymerase II subunit RPB3

• Macromolecule: Name: DNA-directed RNA polymerase II subunit RPB3 / type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO
• Source (natural): Organism: Sus scrofa (pig)
• Molecular weight: Theoretical: 31.439074 KDa

Sequence

String:

MPYANQPTVR ITELTDENVK FIIENTDLAV ANSIRRVFIA EVPIIAIDWV QIDANSSVLH DEFIAHRLGL IPLTSDDIVD KLQYSRDCT CEEFCPECSV EFTLDVRCNE DQTRHVTSRD LISNSPRVIP VTSRNRDNDP SDYVEQDDIL IVKLRKGQEL R LRAYAKKG FGKEHAKWNP TAGVAFEYDP DNALRHTVYP KPEEWPKSEY SELDEDESQA PYDPNGKPER FYYNVESCGS LR PETIVLS ALSGLKKKLS DLQTQLSHEI QSDVLTIN

UniProtKB: DNA-directed RNA polymerase II subunit RPB3

Macromolecule #4: RNA polymerase II subunit D

• Macromolecule: Name: RNA polymerase II subunit D / type: protein_or_peptide / ID: 4 / Number of copies: 1 / Enantiomer: LEVO
• Source (natural): Organism: Sus scrofa (pig)
• Molecular weight: Theoretical: 20.962621 KDa

Sequence

String:

MWGPAQPYSD SALSPKPRPF RAVFRGSALP FPAVRVEVRG RSMAAGGSDP RAGDVEEDAS QLIFPKEFET AETLLNSEVH MLLEHRKQQ NESAEDEQEL SEVFMKTLNY TARFSRFKNR ETIASVRSLL LQKKLHKFEL ACLANLCPET AEESKALIPS L EGRFEDEE LQQILDDIQT KRSFQY

UniProtKB: RNA polymerase II subunit D

Macromolecule #5: DNA-directed RNA polymerase II subunit E

• Macromolecule: Name: DNA-directed RNA polymerase II subunit E / type: protein_or_peptide / ID: 5 / Number of copies: 1 / Enantiomer: LEVO
• Source (natural): Organism: Sus scrofa (pig)
• Molecular weight: Theoretical: 24.644318 KDa

Sequence

String:

MDDEEETYRL WKIRKTIMQL CHDRGYLVTQ DELDQTLEEF KAQFGDKPSE GRPRRTDLTV LVAHNDDPTD QMFVFFPEEP KVGIKTIKV YCQRMQEENI TRALIVVQQG MTPSAKQSLV DMAPKYILEQ FLQQELLINI TEHELVPEHV VMTKEEVTEL L ARYKLREN QLPRIQAGDP VARYFGIKRG QVVKIIRPSE TAGRYITYRL VQ

UniProtKB: DNA-directed RNA polymerase II subunit E

Macromolecule #6: DNA-directed RNA polymerase II subunit F

• Macromolecule: Name: DNA-directed RNA polymerase II subunit F / type: protein_or_peptide / ID: 6 / Number of copies: 1 / Enantiomer: LEVO
• Source (natural): Organism: Sus scrofa (pig)
• Molecular weight: Theoretical: 14.477001 KDa

Sequence

String:

MSDNEDNFDG DDFDDVEEDE GLDDLENAEE EGQENVEILP SGERPQANQK RITTPYMTKY ERARVLGTRA LQIAMCAPVM VELEGETDP LLIAMKELKA RKIPIIIRRY LPDGSYEDWG VDELIISD

UniProtKB: DNA-directed RNA polymerases I, II, and III subunit RPABC2

Macromolecule #7: DNA-directed RNA polymerase II subunit RPB7

• Macromolecule: Name: DNA-directed RNA polymerase II subunit RPB7 / type: protein_or_peptide / ID: 7 / Number of copies: 1 / Enantiomer: LEVO
• Source (natural): Organism: Sus scrofa (pig)
• Molecular weight: Theoretical: 19.314283 KDa

Sequence

String:

MFYHISLEHE ILLHPRYFGP NLLNTVKQKL FTEVEGTCTG KYGFVIAVTT IDNIGAGVIQ PGRGFVLYPV KYKAIVFRPF KGEVVDAVV TQVNKVGLFT EIGPMSCFIS RHSIPSEMEF DPNSNPPCYK TMDEDIVIQQ DDEIRLKIVG TRVDKNDIFA I GSLMDDYL GLVS

UniProtKB: DNA-directed RNA polymerase II subunit RPB7

Macromolecule #8: DNA-directed RNA polymerases I, II, and III subunit RPABC3

• Macromolecule: Name: DNA-directed RNA polymerases I, II, and III subunit RPABC3; type: protein_or_peptide / ID: 8 / Number of copies: 1 / Enantiomer: LEVO
• Source (natural): Organism: Sus scrofa (pig)
• Molecular weight: Theoretical: 17.162273 KDa

Sequence

String:

MAGILFEDIF DVKDIDPEGK KFDRVSRLHC ESESFKMDLI LDVNIQIYPV DLGDKFRLVI ASTLYEDGTL DDGEYNPTDD RPSRADQFE YVMYGKVYRI EGDETSTEAA TRLSAYVSYG GLLMRLQGDA NNLHGFEVDS RVYLLMKKLA F

UniProtKB: DNA-directed RNA polymerases I, II, and III subunit RPABC3

Macromolecule #9: DNA-directed RNA polymerase II subunit RPB9

• Macromolecule: Name: DNA-directed RNA polymerase II subunit RPB9 / type: protein_or_peptide / ID: 9 / Number of copies: 1 / Enantiomer: LEVO
• Source (natural): Organism: Sus scrofa (pig)
• Molecular weight: Theoretical: 14.541221 KDa

Sequence

String:

MEPDGTYEPG FVGIRFCQEC NNMLYPKEDK ENRILLYACR NCDYQQEADN SCIYVNKITH EVDELTQIIA DVSQDPTLPR TEDHPCQKC GHKEAVFFQS HSARAEDAMR LYYVCTAPHC GHRWTE

UniProtKB: DNA-directed RNA polymerase II subunit RPB9

Macromolecule #10: DNA-directed RNA polymerases I, II, and III subunit RPABC5

• Macromolecule: Name: DNA-directed RNA polymerases I, II, and III subunit RPABC5; type: protein_or_peptide / ID: 10 / Number of copies: 1 / Enantiomer: LEVO
• Source (natural): Organism: Sus scrofa (pig)
• Molecular weight: Theoretical: 7.655123 KDa

Sequence

String:

MIIPVRCFTC GKIVGNKWEA YLGLLQAEYT EGDALDALGL KRYCCRRMLL AHVDLIEKLL NYAPLEK

UniProtKB: DNA-directed RNA polymerases I, II, and III subunit RPABC5

Macromolecule #11: RNA_pol_L_2 domain-containing protein

• Macromolecule: Name: RNA_pol_L_2 domain-containing protein / type: protein_or_peptide / ID: 11 / Number of copies: 1 / Enantiomer: LEVO
• Source (natural): Organism: Sus scrofa (pig)
• Molecular weight: Theoretical: 13.310284 KDa

Sequence

String:

MNAPPAFESF LLFEGEKKIT INKDTKVPNA CLFTINKEDH TLGNIIKSQL LKDPQVLFAG YKVPHPLEHK IIIRVQTTPD YSPQEAFTN AITDLISELS LLEERFRVAI KDKQEGIE

UniProtKB: DNA-directed RNA polymerase II subunit RPB11-a

Macromolecule #12: RNA polymerase II subunit K

• Macromolecule: Name: RNA polymerase II subunit K / type: protein_or_peptide / ID: 12 / Number of copies: 1 / Enantiomer: LEVO
• Source (natural): Organism: Sus scrofa (pig)
• Molecular weight: Theoretical: 7.018244 KDa

Sequence

String:

MDTQKDVQPP KQQPMIYICG ECHTENEIKS RDPIRCRECG YRIMYKKRTK RLVVFDAR

UniProtKB: RNA polymerase II, I and III subunit K

Macromolecule #13: Transcription elongation factor SPT5

• Macromolecule: Name: Transcription elongation factor SPT5 / type: protein_or_peptide / ID: 13 / Number of copies: 1 / Enantiomer: LEVO
• Source (natural): Organism: Homo sapiens (human)
• Molecular weight: Theoretical: 121.145477 KDa
• Recombinant expression: Organism: Escherichia coli BL21(DE3) (bacteria)

Sequence

String:

MSDSEDSNFS EEEDSERSSD GEEAEVDEER RSAAGSEKEE EPEDEEEEEE EEEYDEEEEE EDDDRPPKKP RHGGFILDEA DVDDEYEDE DQWEDGAEDI LEKEEIEASN IDNVVLDEDR SGARRLQNLW RDQREEELGE YYMKKYAKSS VGETVYGGSD E LSDDITQQ QLLPGVKDPN LWTVKCKIGE ERATAISLMR KFIAYQFTDT PLQIKSVVAP EHVKGYIYVE AYKQTHVKQA IE GVGNLRL GYWNQQMVPI KEMTDVLKVV KEVANLKPKS WVRLKRGIYK DDIAQVDYVE PSQNTISLKM IPRIDYDRIK ARM SLKDWF AKRKKFKRPP QRLFDAEKIR SLGGDVASDG DFLIFEGNRY SRKGFLFKSF AMSAVITEGV KPTLSELEKF EDQP EGIDL EVVTESTGKE REHNFQPGDN VEVCEGELIN LQGKILSVDG NKITIMPKHE DLKDMLEFPA QELRKYFKMG DHVKV IAGR FEGDTGLIVR VEENFVILFS DLTMHELKVL PRDLQLCSET ASGVDVGGQH EWGELVQLDP QTVGVIVRLE RETFQV LNM YGKVVTVRHQ AVTRKKDNRF AVALDSEQNN IHVKDIVKVI DGPHSGREGE IRHLFRSFAF LHCKKLVENG GMFVCKT RH LVLAGGSKPR DVTNFTVGGF APMSPRISSP MHPSAGGQRG GFGSPGGGSG GMSRGRGRRD NELIGQTVRI SQGPYKGY I GVVKDATEST ARVELHSTCQ TISVDRQRLT TVGSRRPGGM TSTYGRTPMY GSQTPMYGSG SRTPMYGSQT PLQDGSRTP HYGSQTPLHD GSRTPAQSGA WDPNNPNTPS RAEEEYEYAF DDEPTPSPQA YGGTPNPQTP GYPDPSSPQV NPQYNPQTPG TPAMYNTDQ FSPYAAPSPQ GSYQPSPSPQ SYHQVAPSPA GYQNTHSPAS YHPTPSPMAY QASPSPSPVG YSPMTPGAPS P GGYNPHTP GSGIEQNSSD WVTTDIQVKV RDTYLDTQVV GQTGVIRSVT GGMCSVYLKD SEKVVSISSE HLEPITPTKN NK VKVILGE DREATGVLLS IDGEDGIVRM DLDEQLKILN LRFLGKLLEA

UniProtKB: Transcription elongation factor SPT5

Macromolecule #14: RNA

• Macromolecule: Name: RNA / type: rna / ID: 14 ; Details: The last nucleotide 47 (A) in the sample was extended to the 3' of the original scaffolding RNA (46-mer) during sample preparation in the presence of ATP.; Number of copies: 1
• Source (natural): Organism: HIV whole-genome vector AA1305#18 (others)
• Molecular weight: Theoretical: 15.076017 KDa

Sequence

String:

ACCAGAUCUG AGCCUGGGAG CUCUCUGGCU AACUAGGGAA CCCACUA

Macromolecule #15: Non-template DNA

• Macromolecule: Name: Non-template DNA / type: dna / ID: 15 / Number of copies: 1 / Classification: DNA
• Source (natural): Organism: HIV whole-genome vector AA1305#18 (others)
• Molecular weight: Theoretical: 14.672441 KDa

Sequence

String:

(DG)(DT)(DA)(DC)(DA)(DC)(DT)(DA)(DG)(DT) (DA)(DC)(DC)(DT)(DA)(DC)(DT)(DC)(DG)(DA) (DG)(DT)(DG)(DA)(DC)(DC)(DT)(DT)(DA) (DA)(DG)(DC)(DC)(DT)(DC)(DA)(DA)(DT)(DA) (DA) (DA)(DG)(DC)(DT)(DT)(DG)(DC)(DC)

Macromolecule #16: Template DNA

• Macromolecule: Name: Template DNA / type: dna / ID: 16 / Number of copies: 1 / Classification: DNA
• Source (natural): Organism: HIV whole-genome vector AA1305#18 (others)
• Molecular weight: Theoretical: 14.93458 KDa

Sequence

String:

(DG)(DG)(DC)(DA)(DA)(DG)(DC)(DT)(DT)(DT) (DA)(DT)(DT)(DG)(DA)(DG)(DG)(DC)(DT)(DT) (DA)(DA)(DG)(DC)(DA)(DG)(DT)(DG)(DG) (DG)(DT)(DT)(DC)(DA)(DA)(DG)(DG)(DT)(DA) (DC) (DT)(DA)(DG)(DT)(DG)(DT)(DA)(DC)

Macromolecule #17: MAGNESIUM ION

• Macromolecule: Name: MAGNESIUM ION / type: ligand / ID: 17 / Number of copies: 1 / Formula: MG
• Molecular weight: Theoretical: 24.305 Da

Macromolecule #18: ZINC ION

• Macromolecule: Name: ZINC ION / type: ligand / ID: 18 / Number of copies: 8 / Formula: ZN
• Molecular weight: Theoretical: 65.409 Da

Experimental details

Structure determination

• Method: cryo EM
• Processing: single particle reconstruction
• Aggregation state: particle

Sample preparation

• Buffer: pH: 7.4
• Vitrification: Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277.15 K / Instrument: FEI VITROBOT MARK IV

Electron microscopy

• Microscope: FEI TITAN KRIOS
• Image recording: Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Detector mode: COUNTING / Average electron dose: 43.21 e/Ų
• Electron beam: Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
• Electron optics: Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD
• Experimental equipment: Model: Titan Krios / Image courtesy: FEI Company

Image processing

• Startup model: Type of model: OTHER / Details: Ab initio model created by CryoSPARC
• Final reconstruction: Resolution.type: BY AUTHOR / Resolution: 3.0 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 194785
• Initial angle assignment: Type: MAXIMUM LIKELIHOOD
• Final angle assignment: Type: MAXIMUM LIKELIHOOD

Atomic model buiding 1

• Refinement: Space: REAL

Output model

PDB-7ol0:
Structure of active transcription elongation complex Pol II-DSIF (SPT5-KOW5)