EMDB-48623: Structure of a native Drosophila melanogaster Pol II Elongation C...

Basic information

Entry

Database: EMDB / ID: EMD-48623

• Title: Structure of a native Drosophila melanogaster Pol II Elongation Complex without Rpb4/Rpb7 stalk
• Map data: Refined, unsharpened Coulomb potential density map. Particles extracted in 440x440 Fourier cropped to 360x360.

Sample

• Complex: Native purified Pol II elongation complex from Drosophila melanogaster without the Rpb4/Rpb7 stalk
  • Protein or peptide: x 4 types
  • RNA: x 1 types
  • DNA: x 2 types
• Protein or peptide: x 6 types
• Ligand: x 1 types

• Keywords: polymerase / Pol II / transcription / mRNA

Function / homology

Function:

Formation of the Early Elongation Complex / mRNA Capping / mRNA Splicing - Minor Pathway / RNA Polymerase I Transcription Initiation / RNA Polymerase III Transcription Initiation From Type 1 Promoter / RNA Polymerase III Transcription Initiation From Type 2 Promoter / RNA Pol II CTD phosphorylation and interaction with CE / Formation of RNA Pol II elongation complex / TP53 Regulates Transcription of DNA Repair Genes / RNA Polymerase II Transcription Elongation / RNA Polymerase II Promoter Escape / RNA Polymerase II Transcription Pre-Initiation And Promoter Opening / RNA Polymerase II Transcription Initiation / RNA Polymerase II Transcription Initiation And Promoter Clearance / RNA Polymerase II Pre-transcription Events / Gap-filling DNA repair synthesis and ligation in TC-NER / Formation of TC-NER Pre-Incision Complex / Processing of Capped Intron-Containing Pre-mRNA / Dual incision in TC-NER / polytene chromosome puff / RNA polymerase II transcribes snRNA genes / mRNA Splicing - Major Pathway / RNA Polymerase I Promoter Escape / Transcriptional regulation by small RNAs / Estrogen-dependent gene expression / polytene chromosome / maintenance of transcriptional fidelity during transcription elongation by RNA polymerase II / transcription by RNA polymerase I / RNA polymerase I complex / transcription elongation by RNA polymerase I / RNA polymerase III complex / transcription-coupled nucleotide-excision repair / RNA polymerase II, core complex / tRNA transcription by RNA polymerase III / : / DNA-directed RNA polymerase activity / transcription initiation at RNA polymerase II promoter / ribonucleoside binding / : / : / : / : / : / : / DNA-directed RNA polymerase / mitotic cell cycle / cellular response to heat / transcription by RNA polymerase II / nucleic acid binding / protein dimerization activity / nucleolus / DNA binding / zinc ion binding / metal ion binding / nucleus / cytoplasm

Domain/homology:

RNA polymerase II, heptapeptide repeat, eukaryotic / Eukaryotic RNA polymerase II heptapeptide repeat. / RNA polymerase Rpb1, domain 6 / RNA polymerase Rpb1, domain 6 / RNA polymerase Rpb1, domain 7 / RNA polymerase Rpb1, domain 7 superfamily / RNA polymerase Rpb1, domain 7 / Pol II subunit B9, C-terminal zinc ribbon / RNA polymerase RBP11 / Zinc finger TFIIS-type signature. / RNA polymerase Rpb2, domain 5 / RNA polymerase Rpb2, domain 5 / RNA polymerase Rpb2, domain 4 / RNA polymerase Rpb2, domain 4 / DNA-directed RNA polymerase, M/15kDa subunit / RNA polymerases M/15 Kd subunit / RNA polymerase subunit 9 / 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 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, 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 / Transcription factor S-II (TFIIS) / Zinc finger TFIIS-type profile. / C2C2 Zinc finger / RNA polymerase subunit RPABC4/transcription elongation factor Spt4 / DNA-directed RNA polymerase, RBP11-like dimerisation domain / RNA polymerase Rpb3/Rpb11 dimerisation domain / RNA polymerase, subunit H/Rpb5 C-terminal / DNA-directed RNA polymerase subunit Rpo5/Rpb5 / 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. / DNA-directed RNA polymerase, subunit beta-prime / RNA polymerase Rpb6 / RNA polymerase, subunit omega/Rpo6/RPB6 / RNA polymerase Rpb6 / RNA polymerase Rpb2, domain 2 superfamily / RNA polymerase Rpb1, domain 3 superfamily / RPB6/omega subunit-like superfamily / 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 / RNA polymerase Rpb1, clamp domain superfamily / RNA polymerase Rpb1, domain 3 / RNA polymerase Rpb1, domain 3 / RNA polymerase Rpb1, domain 1 / RNA polymerase Rpb1, domain 1 / RNA polymerase, beta subunit, protrusion / RNA polymerase beta subunit / RNA polymerase, alpha subunit / RNA polymerase Rpb1, domain 5 / RNA polymerase Rpb1, domain 4 / RNA polymerase Rpb1, domain 2 / RNA polymerase Rpb1, domain 5 / RNA polymerase Rpb1, domain 4 / DNA-directed RNA polymerase, insert domain superfamily / RNA polymerase, N-terminal / RNA polymerase Rpb1, funnel domain superfamily / RNA polymerase I subunit A N-terminus / RNA polymerase Rpb2, domain 2 / RNA polymerase Rpb2, domain 2 / RNA polymerase, RBP11-like subunit / RNA polymerase, beta subunit, conserved site / RNA polymerase Rpb2, domain 7 / RNA polymerase Rpb2, domain 3 / RNA polymerase Rpb2, OB-fold / RNA polymerase Rpb2, domain 7 / RNA polymerase Rpb2, domain 3 / RNA polymerases beta chain signature. / DNA-directed RNA polymerase, subunit 2, hybrid-binding domain / DNA-directed RNA polymerase, subunit 2

Component:

GH07456p / DNA-directed RNA polymerase II subunit RPB1 / DNA-directed RNA polymerase II subunit RPB2 / DNA-directed RNA polymerase II subunit RPB9 / DNA-directed RNA polymerases I, II, and III subunit RPABC2 / GEO11084p1 / RPB5 homolog / DNA-directed RNA polymerases I, II, and III subunit RPABC5 / DNA-directed RNA polymerase II subunit RPB11 / DNA-directed RNA polymerases I, II, and III subunit RPABC3

• Biological species: Drosophila melanogaster (fruit fly)
• Method: single particle reconstruction / cryo EM / Resolution: 3.37 Å
• Authors: Venette-Smith NL / Vishwakarma RK / Dollinger R / Schultz J / Venkatakrishnan V / Babitzke P / Anand G / Gilmour DS / Armache J-P / Murakami K

Funding support

United States, 3 items

Organization	Grant number	Country
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)	GM131860	United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)	GM047477	United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)	GM098399	United States

• Citation: Journal: To Be Published; Title: Structural Characterization of Native RNA Polymerase II Transcription Complexes in Drosophila melanogaster; Authors: Venette-Smith NL / Vishwakarma RK / Dollinger R / Schultz J / Venkatakrishnan V / Babitzke P / Anand G / Gilmour DS / Armache J-P / Murakami K

History

Deposition	Jan 13, 2025	-
Header (metadata) release	May 14, 2025	-
Map release	May 14, 2025	-
Update	May 14, 2025	-
Current status	May 14, 2025	Processing site: RCSB / Status: Released

Map

• File: Download / File: emd_48623.map.gz / Format: CCP4 / Size: 178 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
• Annotation: Refined, unsharpened Coulomb potential density map. Particles extracted in 440x440 Fourier cropped to 360x360.
• Voxel size: X=Y=Z: 1.1538 Å

Density

Contour Level	By AUTHOR: 0.4
Minimum - Maximum	-0.0 - 1.5666236
Average (Standard dev.)	0.0065101567 (±0.04348934)

• Symmetry: Space group: 1

Details

EMDB XML:

Map geometry	Axis order X Y Z Origin 0 0 0 Dimensions 360 360 360 Spacing 360 360 360
Cell	A=B=C: 415.368 Å α=β=γ: 90.0 °

Supplemental data

Additional map: Refined, auto-sharpened Coulomb potential density map with Bfactor...

• File: emd_48623_additional_1.map
• Annotation: Refined, auto-sharpened Coulomb potential density map with Bfactor = -51.7

Half map: Half-map 2

• File: emd_48623_half_map_1.map
• Annotation: Half-map 2

Half map: Half-map 1

• File: emd_48623_half_map_2.map
• Annotation: Half-map 1

Sample components

Entire : Native purified Pol II elongation complex from Drosophila melanog...

• Entire: Name: Native purified Pol II elongation complex from Drosophila melanogaster without the Rpb4/Rpb7 stalk

Components

• Complex: Native purified Pol II elongation complex from Drosophila melanogaster without the Rpb4/Rpb7 stalk
  • Protein or peptide: DNA-directed RNA polymerases I, II, and III subunit RPABC2
  • Protein or peptide: DNA-directed RNA polymerases I, II, and III subunit RPABC3
  • Protein or peptide: DNA-directed RNA polymerase II subunit RPB11
  • RNA: RNA
  • DNA: Template DNA
  • DNA: Non-template DNA
  • Protein or peptide: RPB5 homolog
• Protein or peptide: DNA-directed RNA polymerase II subunit RPB1
• Protein or peptide: DNA-directed RNA polymerase II subunit RPB2
• Protein or peptide: DNA-directed RNA polymerase II subunit RPB3
• 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: DNA-directed RNA polymerase II subunit RPB12
• Ligand: ZINC ION

Supramolecule #1: Native purified Pol II elongation complex from Drosophila melanog...

• Supramolecule: Name: Native purified Pol II elongation complex from Drosophila melanogaster without the Rpb4/Rpb7 stalk; type: complex / ID: 1 / Parent: 0 / Macromolecule list: #4-#5, #8, #10-#13; Details: This entry represents a native Pol II elongation complex from Drosophila melanogaster without the Rpb4/Rpb7 stalk, obtained through stringent 3D classification and refinement.
• Source (natural): Organism: Drosophila melanogaster (fruit fly)
• Molecular weight: Theoretical: 930 KDa

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 / EC number: DNA-directed RNA polymerase
• Source (natural): Organism: Drosophila melanogaster (fruit fly)
• Molecular weight: Theoretical: 166.420375 KDa

Sequence

String:

SKAPLRQVKR VQFGILSPDE IRRMSVTEGG VQFAETMEGG RPKLGGLMDP RQGVIDRTSR CQTCAGNMTE CPGHFGHIDL AKPVFHIGF ITKTIKILRC VCFYCSKMLV SPHNPKIKEI VMKSRGQPRK RLAYVYDLCK GKTICEGGED MDLTKENQQP D PNKKPGHG GCGHYQPSIR RTGLDLTAEW KHQNEDSQEK KIVVSAERVW EILKHITDEE CFILGMDPKY ARPDWMIVTV LP VPPLAVR PAVVMFGAAK NQDDLTHKLS DIIKANNELR KNEASGAAAH VIQENIKMLQ FHVATLVDND MPGMPRAMQK SGK PLKAIK ARLKGKEGRI RGNLMGKRVD FSARTVITPD PNLRIDQVGV PRSIAQNLTF PELVTPFNID RMQELVRRGN SQYP GAKYI VRDNGERIDL RFHPKSSDLH LQCGYKVERH LRDDDLVIFN RQPTLHKMSM MGHRVKVLPW STFRMNLSCT SPYNA DFDG DEMNLHVPQS METRAEVENI HITPRQIITP QANKPVMGIV QDTLTAVRKM TKRDVFITRE QVMNLLMFLP TWDAKM PQP CILKPRPLWT GKQIFSLIIP GNVNMIRTHS THPDEEDEGP YKWISPGDTK VMVEHGELIM GILCKKSLGT SAGSLLH IC FLELGHDIAG RFYGNIQTVI NNWLLFEGHS IGIGDTIADP QTYNEIQQAI KKAKDDVINV IQKAHNMELE PTPGNTLR Q TFENKVNRIL NDARDKTGGS AKKSLTEYNN LKAMVVSGSK GSNINISQVI ACVGQQNVEG KRIPYGFRKR TLPHFIKDD YGPESRGFVE NSYLAGLTPS EFYFHAMGGR EGLIDTAVKT AETGYIQRRL IKAMESVMVN YDGTVRNSVG QLIQLRYGED GLCGELVEF QNMPTVKLSN KSFEKRFKFD WSNERLMKKV FTDDVIKEMT DSSEAIQELE AEWDRLVSDR DSLRQIFPNG E SKVVLPCN LQRMIWNVQK IFHINKRLPT DLSPIRVIKG VKTLLERCVI VTGNDRISKQ ANENATLLFQ CLIRSTLCTK YV SEEFRLS TEAFEWLVGE IETRFQQAQA NPGEMVGALA AQSLGEPATQ MTLNTFHFAG VSSKNVTLGV PRLKEIINIS KKP KAPSLT VFLTGGAARD AEKAKNVLCR LEHTTLRKVT ANTAIYYDPD PQRTVISEDQ EFVNVYYEMP DFDPTRISPW LLRI ELDRK RMTDKKLTME QIAEKINVGF GEDLNCIFND DNADKLVLRI RIMNNEENKF QDEDEAVDKM EDDMFLRCIE ANMLS DMTL QGIEAIGKVY MHLPQTDSKK RIVITETGEF KAIGEWLLET DGTSMMKVLS ERDVDPIRTS SNDICEIFQV LGIEAV RKS VEKEMNAVLQ FYGLYVNYRH LALLCDVMTA KGHLMAITRH GINRQDTGAL MRCSFEETVD VLMDAAAHAE TDPMRGV SE NIIMGQLPKM GTGCFDLLLD AEKCRFGI

UniProtKB: DNA-directed RNA polymerase II subunit RPB1

Macromolecule #2: DNA-directed RNA polymerase II subunit RPB2

• Macromolecule: Name: DNA-directed RNA polymerase II subunit RPB2 / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO / EC number: DNA-directed RNA polymerase
• Source (natural): Organism: Drosophila melanogaster (fruit fly)
• Molecular weight: Theoretical: 132.640062 KDa

Sequence

String:

ENAEEISHEL WQEACWIVIN AYFDEKGLVR QQLDSFDEFI QMSVQRIVED SPAIELQAEA QHTSGEVETP PRFSLKFEQI YLSKPTHWE KDGSPSPMMP NEARLRNLTY SAPLYVDITK TKNVEGLDPV ETQHQKTFIG KIPIMLRSTY CLLSQLTDRD L TELNECPL DPGGYFIING SEKVLIAQEK MATNTVYVFS MKDGKYAFKT EIRSCLEHSS RPTSTLWVNM MARGSQNIKK SA IGQRIIA ILPYIKQEIP IMIVFRALGF VADRDILEHI IYDFDDPEMM EMVKPSLDEA FVVQEQNVAL NFIGARGARP GVT KDKRIK YAKEILQKEM LPHVGVSDFC ETKKAYFLGY MVHRLLLASL GRRELDDRDH YGNKRLDLAG PLLAFLFRGL FKNL MKEVR MYTQKFIDRG KDFNLELAIK TNIITDGLRY SLATGNWGDQ KKAHQARAGV SQVLNRLTFA STLSHLRRVN SPIGR DGKL AKPRQLHNTL WGMLCPAETP EGAAVGLVKN LALMAYISVG SQPSPILEFL EEWSMENLEE IAPSAIADAT KIFVNG CWV GIHRDPEQLM ATLRKLRRQM DIIVSEVSMI RDIRDREIRI YTDAGRICRP LLIVENGSLL LKKTHVEMLK ERDYNNY SW QVLVASGVVE YIDTLEEETV MIAMSPYDLK QDKDYAYCTT YTHCEIHPAM ILGVCASIIP FPDHNQSPRN TYQSAMGK Q AMGVYITNFH VRMDTLAHVL YYPMKPLVTT RSMEYLRFRE LPAGINSIVA ILCYTGYNQE DSVILNASAV ERGFFRSVF YRSYKDSENK RVGDQEENFE KPHRGTCQGM RNAHYDKLDD DGIIAPGIRV SGDDVVIGKT ITLPENDDEL DSNTKRFSKR DASTFLRNS ETGIVDQVML TLNSEGYKFC KIRVRSVRIP QIGDKFASRH GQKGTCGIQY RQEDMAFTCE GLAPDIIINP H AIPSRMTI GHLIECLQGK LGSNKGEIGD ATPFNDAVNV QKISTFLQEY GYHLRGNEVM YNGHTGRKIN AQVFLGPTYY QR LKHMVDD KIHSRARGPV QILVRQPMEG RARDGGLRFG EMERDCQISH GAAQFLRERL FEVSDPYRVH ICNFCGLIAI ANL RNNTFE CKGCKNKTQI SQVRLPYAAK LLFQELMSMN IAPRLMVT

UniProtKB: DNA-directed RNA polymerase II subunit RPB2

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 / EC number: DNA-directed RNA polymerase
• Source (natural): Organism: Drosophila melanogaster (fruit fly)
• Molecular weight: Theoretical: 30.676199 KDa

Sequence

String:

PYANQPSVQI TELTDDNVKF VLEDTELSVA NSLRRVFIAE TPTLAIDWVQ LEANSTVLSD EFLAHRIGLI PLISDDVVER LQYTRDCIC LDFCPECSVE FTLDVKCSEE QTRHVTTADL KSSNAKVLPV TSRNQGEEDN EYGESNDEIL IIKLRKGQEL K LRAYAKKG FGKEHAKWNP TAGVCFEYDP DNSMRHTLYP KPDEWPKSEH TELEDDQYEA PYNWEAKPNK FFFNVESAGA LK PENIVVM GVQVLKNKLS NLQTQLSHES QN

UniProtKB: GH07456p

Macromolecule #4: DNA-directed RNA polymerases I, II, and III subunit RPABC2

• Macromolecule: Name: DNA-directed RNA polymerases I, II, and III subunit RPABC2; type: protein_or_peptide / ID: 4 / Number of copies: 1 / Enantiomer: LEVO
• Source (natural): Organism: Drosophila melanogaster (fruit fly)
• Molecular weight: Theoretical: 9.73651 KDa

Sequence

String:

GGVPKSKRIT TKYMTKYERA RVLGTRALQI AMCAPIMVEL DGETDPLQIA MKELKQKKIP IIIRRYLPDH SYEDWSIDEL IMVD

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

Macromolecule #5: 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: 5 / Number of copies: 1 / Enantiomer: LEVO
• Source (natural): Organism: Drosophila melanogaster (fruit fly)
• Molecular weight: Theoretical: 16.946137 KDa

Sequence

String:

AGVLFEDIFN VKDMDPEGKK FDRVSRLHCE SESFKMDLIL DINSWLYPME LGDKFRLVLA TTLREDGCPD SGEYNPMEHE GTRADSFEY VMYGKIYRIE GDEAHNEASR LSAYVSFGGL LMRLQGDANN LHGFEVDQHM YLLMKRLA

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

Macromolecule #6: DNA-directed RNA polymerase II subunit RPB9

• Macromolecule: Name: DNA-directed RNA polymerase II subunit RPB9 / type: protein_or_peptide / ID: 6 / Number of copies: 1 / Enantiomer: LEVO
• Source (natural): Organism: Drosophila melanogaster (fruit fly)
• Molecular weight: Theoretical: 13.786586 KDa

Sequence

String:

GFVGIRFCQE CNNMLYPKED KENKILLYAC RNCDYKQEAD SNCIYVNKIM HEIDELTHIV PDVISDPTLP RTEDHACPKC SHREAVFFQ AQTRRAEEEM RLYYVCTNQN CTHRWTE

UniProtKB: DNA-directed RNA polymerase II subunit RPB9

Macromolecule #7: 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: 7 / Number of copies: 1 / Enantiomer: LEVO
• Source (natural): Organism: Drosophila melanogaster (fruit fly)
• Molecular weight: Theoretical: 7.671124 KDa

Sequence

String:

MIIPIRCFTC GKVIGNKWES YLGLLQAEYT EGDALDALGL KRYCCRRMLL GHVDLIEKLL NYAPLEK

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

Macromolecule #8: DNA-directed RNA polymerase II subunit RPB11

• Macromolecule: Name: DNA-directed RNA polymerase II subunit RPB11 / type: protein_or_peptide / ID: 8 / Number of copies: 1 / Enantiomer: LEVO
• Source (natural): Organism: Drosophila melanogaster (fruit fly)
• Molecular weight: Theoretical: 13.28824 KDa

Sequence

String:

MNAPPTFESF LLYEGEKKII KELDTKVTNA AIFTINKEDH TLGNMIRNQL LKDPNVLFAG YKVPHPLEHK FVIRIQTTAD YSPQEAFMN AITDLLAELS LFEERFKDAI KEKKEGG

UniProtKB: DNA-directed RNA polymerase II subunit RPB11

Macromolecule #9: DNA-directed RNA polymerase II subunit RPB12

• Macromolecule: Name: DNA-directed RNA polymerase II subunit RPB12 / type: protein_or_peptide / ID: 9 / Number of copies: 1 / Enantiomer: LEVO
• Source (natural): Organism: Drosophila melanogaster (fruit fly)
• Molecular weight: Theoretical: 5.647678 KDa

Sequence

String:

TAMTYICGEC HHENEMRPRD PIRCRECGYR IMYKKRTKRL VVFDAR

UniProtKB: GEO11084p1

Macromolecule #13: RPB5 homolog

• Macromolecule: Name: RPB5 homolog / type: protein_or_peptide / ID: 13 / Number of copies: 1 / Enantiomer: LEVO
• Source (natural): Organism: Drosophila melanogaster (fruit fly)
• Molecular weight: Theoretical: 24.518332 KDa

Sequence

String:

MDDEAETYKL WRIRKTIMQL SHDRGYLVTQ DELDQTLEQF KEMFGDKPSE KRPARSDLIV LVAHNDDPTD QMFVFFPEEP KIGIKTIKT YCTRMQEENI HRAIVVVQGG MTPSAKQSLV DMAPKYILEQ FLESELLINI TEHELVPEHV VMTVEEKQEL L SRYKLKEN MLMRIQAGDP VARYFGLKRG QVVKIIRSSE TAGRYISYRL VC

UniProtKB: RPB5 homolog

Macromolecule #10: RNA

• Macromolecule: Name: RNA / type: rna / ID: 10 / Number of copies: 1
• Source (natural): Organism: Drosophila melanogaster (fruit fly)
• Molecular weight: Theoretical: 3.2471 KDa

Sequence

String:

AAAAAAAAAA

Macromolecule #11: Template DNA

• Macromolecule: Name: Template DNA / type: dna / ID: 11 / Number of copies: 1 / Classification: DNA
• Source (natural): Organism: Drosophila melanogaster (fruit fly)
• Molecular weight: Theoretical: 15.134661 KDa

Sequence

String:

(DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT) (DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT) (DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT) (DT)(DT)(DT)(DC)(DC)(DT)(DT)(DT)(DT)(DT) (DT) (DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT) (DT)(DT)

Macromolecule #12: Non-template DNA

• Macromolecule: Name: Non-template DNA / type: dna / ID: 12 / Number of copies: 1 / Classification: DNA
• Source (natural): Organism: Drosophila melanogaster (fruit fly)
• Molecular weight: Theoretical: 12.170105 KDa

Sequence

String:

(DA)(DA)(DA)(DA)(DA)(DA)(DA)(DA)(DA)(DA) (DA)(DA)(DA)(DA)(DA)(DA)(DA)(DA)(DA)(DA) (DA)(DA)(DA)(DA)(DA)(DA)(DA)(DA)(DA) (DA)(DA)(DA)(DA)(DA)(DA)(DA)(DA)(DA)(DA)

Macromolecule #14: ZINC ION

• Macromolecule: Name: ZINC ION / type: ligand / ID: 14 / 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

• Concentration: 1.0 mg/mL

Buffer

pH: 7.5
Component:

Concentration	Formula	Name
150.0 mM	NaCl	Sodium chloride
10.0 mM	HEPES-HCl	HEPES
5.0 percent	Glycerol	Glycerol
1.0 mM	EDTA	EDTA
350.0 ug/mL	FLAG	FLAG peptide

Details: 10 mM HEPES-HCl (pH = 7.5), 150 mM NaCl, 5% glycerol, 1 mM EDTA, 350 ug/mL 3x FLAG peptide, 1/1000th protease inhibitor

• Grid: Model: Quantifoil / Material: GOLD / Mesh: 200 / Support film - Material: GOLD / Support film - topology: HOLEY
• Vitrification: Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 4 K / Instrument: FEI VITROBOT MARK IV
• Details: double FLAG-tagged Pol II subunit Rpb1 was used for purification of Pol II complexes

Electron microscopy

• Microscope: TFS TALOS
• Image recording: Film or detector model: FEI FALCON IV (4k x 4k) / Digitization - Dimensions - Width: 4096 pixel / Digitization - Dimensions - Height: 4096 pixel / Number grids imaged: 4 / Number real images: 31103 / Average electron dose: 50.65 e/Ų; Details: Although the data was collected, motion-corrected and CTF estimated using the pixel size of 0.944, all the subsequent data processing was performed using pixel size of 1.1538. This was achieved by extracting particles in box size 440x440 and Fourier-cropping it to box size 360x360
• Electron beam: Acceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
• Electron optics: Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.5 µm / Nominal defocus min: 1.0 µm
• Sample stage: Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN

Image processing

• Particle selection: Number selected: 2349308 ; Details: Initially, nearly 15 million particles were picked from micrographs. However, the number of particles cited here (2,349,308) is the starting number of particles after initial cleanup and 3D classification of the data.
• CTF correction: Software - Name: cryoSPARC (ver. 4.3.1) / Details: 'Patch CTF Estimation' in cryoSPARC / Type: PHASE FLIPPING AND AMPLITUDE CORRECTION
• Startup model: Type of model: INSILICO MODEL / In silico model: 'Ab initio' function in cryoSPARC
• Final reconstruction: Number classes used: 1 / Applied symmetry - Point group: C₁ (asymmetric) / Algorithm: BACK PROJECTION / Resolution.type: BY AUTHOR / Resolution: 3.37 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 4.3.1) / Number images used: 96409
• Initial angle assignment: Type: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 4.3.1)
• Final angle assignment: Type: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 4.3.1) / Details: 'Non-Uniform Refinement' in cryoSPARC

Atomic model buiding 1

Initial model

PDB ID	Chain
4052	chain_id: A, source_name: AlphaFold, initial_model_type: in silico model
8266	chain_id: B, source_name: AlphaFold, initial_model_type: in silico model
7183	chain_id: C, source_name: AlphaFold, initial_model_type: in silico model
vea5	chain_id: D, source_name: AlphaFold, initial_model_type: in silico model
jzf5	chain_id: E, source_name: AlphaFold, initial_model_type: in silico model
4320	chain_id: F, source_name: AlphaFold, initial_model_type: in silico model
vfb5	chain_id: G, source_name: AlphaFold, initial_model_type: in silico model
vnz3	chain_id: H, source_name: AlphaFold, initial_model_type: in silico model
6958	chain_id: I, source_name: AlphaFold, initial_model_type: in silico model
vc49	chain_id: J, source_name: AlphaFold, initial_model_type: in silico model
vje4	chain_id: K, source_name: AlphaFold, initial_model_type: in silico model
ige3	chain_id: L, source_name: AlphaFold, initial_model_type: in silico model

• Details: The model was based on PDB: 6GML. First, 6GML was rigid-body fit in the cryo-EM Coulomb potential density map using UCSF ChimeraX. Next, AlphaFold2 models for individual Drosophila melanogaster Pol II subunits were aligned to their 6GML counterparts, and further optimized in the density using rigid-body fit. For the final model optimization, phenix.real_space_refine was used
• Refinement: Protocol: AB INITIO MODEL / Overall B value: 50

Output model

PDB-9mu8:
Structure of a native Drosophila melanogaster Pol II Elongation Complex without Rpb4/Rpb7 stalk