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- EMDB-23895: Cryo-EM structure of Escherichia coli RNA polymerase bound to lam... -

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Basic information

Entry
Database: EMDB / ID: EMD-23895
TitleCryo-EM structure of Escherichia coli RNA polymerase bound to lambda PR (-5G to C) promoter DNA
Map data
Sample
  • Complex: Escherichia coli RNA polymerase bound to lambda PR promoter DNA (-5G to C)
    • Complex: RNA polymerase
      • Protein or peptide: DNA-directed RNA polymerase subunit alphaPolymerase
      • Protein or peptide: DNA-directed RNA polymerase subunit betaPolymerase
      • Protein or peptide: DNA-directed RNA polymerase subunit beta'Polymerase
      • Protein or peptide: DNA-directed RNA polymerase subunit omegaPolymerase
      • Protein or peptide: RNA polymerase sigma factor RpoD
    • Complex: lambda PR (-5G to C) promoter DNA
      • DNA: Nontemplate strand of lambda PR DNA promoter (-5G to C)
      • DNA: Template strand of lambda PR promoter DNA (-5C to G)
  • Ligand: CHAPSOCHAPS detergent
  • Ligand: MAGNESIUM ION
  • Ligand: ZINC ION
Function / homology
Function and homology information


sigma factor activity / DNA-directed RNA polymerase complex / DNA-templated transcription initiation / ribonucleoside binding / DNA-directed 5'-3' RNA polymerase activity / DNA-directed RNA polymerase / protein dimerization activity / DNA-templated transcription / magnesium ion binding / DNA binding ...sigma factor activity / DNA-directed RNA polymerase complex / DNA-templated transcription initiation / ribonucleoside binding / DNA-directed 5'-3' RNA polymerase activity / DNA-directed RNA polymerase / protein dimerization activity / DNA-templated transcription / magnesium ion binding / DNA binding / zinc ion binding / metal ion binding / cytoplasm
Similarity search - Function
RNA polymerase sigma factor 70, non-essential domain / Sigma-70, non-essential region / RNA polymerase sigma factor 70, region 1.1 / Sigma-70 factor, region 1.1 superfamily / Sigma-70 factor, region 1.1 / Sigma-70 factors family signature 1. / RNA polymerase sigma factor RpoD, C-terminal / RNA polymerase sigma factor RpoD / RNA polymerase sigma-70 region 1.2 / Sigma-70 factor, region 1.2 ...RNA polymerase sigma factor 70, non-essential domain / Sigma-70, non-essential region / RNA polymerase sigma factor 70, region 1.1 / Sigma-70 factor, region 1.1 superfamily / Sigma-70 factor, region 1.1 / Sigma-70 factors family signature 1. / RNA polymerase sigma factor RpoD, C-terminal / RNA polymerase sigma factor RpoD / RNA polymerase sigma-70 region 1.2 / Sigma-70 factor, region 1.2 / RNA polymerase sigma-70 region 3 / Sigma-70 region 3 / Sigma-70 factors family signature 2. / RNA polymerase sigma-70 / RNA polymerase sigma-70 region 4 / Sigma-70, region 4 / RNA polymerase sigma-70 region 2 / RNA polymerase sigma-70 like domain / Sigma-70 region 2 / RNA polymerase sigma factor, region 2 / RNA polymerase sigma factor, region 3/4-like / DNA-directed RNA polymerase, omega subunit / DNA-directed RNA polymerase, subunit beta-prime, bacterial type / DNA-directed RNA polymerase, beta subunit, external 1 domain superfamily / DNA-directed RNA polymerase, beta subunit, external 1 domain / RNA polymerase beta subunit external 1 domain / RNA polymerase, alpha subunit, C-terminal / Bacterial RNA polymerase, alpha chain C terminal domain / DNA-directed RNA polymerase, alpha subunit / DNA-directed RNA polymerase beta subunit, bacterial-type / RNA polymerase Rpb6 / RNA polymerase, subunit omega/Rpo6/RPB6 / RNA polymerase Rpb6 / RNA polymerase Rpb1, domain 3 superfamily / RNA polymerase Rpb1, clamp domain superfamily / RPB6/omega subunit-like superfamily / DNA-directed RNA polymerase, subunit beta-prime / RNA polymerase Rpb1, domain 3 / RNA polymerase Rpb1, domain 3 / RNA polymerase Rpb2, domain 2 superfamily / RNA polymerase Rpb1, domain 1 / RNA polymerase Rpb1, domain 1 / RNA polymerase, alpha subunit / RNA polymerase Rpb1, domain 4 / RNA polymerase Rpb1, domain 2 / RNA polymerase Rpb1, domain 4 / RNA polymerase, N-terminal / RNA polymerase Rpb1, funnel domain superfamily / RNA polymerase I subunit A N-terminus / RNA polymerase Rpb1, domain 5 / RNA polymerase Rpb1, domain 5 / 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 / RNA polymerase Rpb2, domain 2 / 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 / DNA-directed RNA polymerase, subunit 2, hybrid-binding domain superfamily / RNA polymerase Rpb2, domain 6 / Winged helix-like DNA-binding domain superfamily
Similarity search - Domain/homology
DNA-directed RNA polymerase subunit alpha / DNA-directed RNA polymerase subunit beta' / DNA-directed RNA polymerase subunit omega / DNA-directed RNA polymerase subunit beta / RNA polymerase sigma factor RpoD
Similarity search - Component
Biological speciesEscherichia coli (E. coli) / Escherichia virus Lambda
Methodsingle particle reconstruction / cryo EM / Resolution: 3.5 Å
AuthorsSaecker RM / Darst SA / Chen J
Funding support United States, 1 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R35 GM118130 United States
CitationJournal: Proc Natl Acad Sci U S A / Year: 2021
Title: Structural origins of RNA polymerase open promoter complex stability.
Authors: Ruth M Saecker / James Chen / Courtney E Chiu / Brandon Malone / Johanna Sotiris / Mark Ebrahim / Laura Y Yen / Edward T Eng / Seth A Darst /
Abstract: The first step in gene expression in all organisms requires opening the DNA duplex to expose one strand for templated RNA synthesis. In , promoter DNA sequence fundamentally determines how fast the ...The first step in gene expression in all organisms requires opening the DNA duplex to expose one strand for templated RNA synthesis. In , promoter DNA sequence fundamentally determines how fast the RNA polymerase (RNAP) forms "open" complexes (RPo), whether RPo persists for seconds or hours, and how quickly RNAP transitions from initiation to elongation. These rates control promoter strength in vivo, but their structural origins remain largely unknown. Here, we use cryoelectron microscopy to determine the structures of RPo formed de novo at three promoters with widely differing lifetimes at 37 °C: λP (t ∼10 h), T7A1 (t ∼4 min), and a point mutant in λP (λP) (t ∼2 h). Two distinct RPo conformers are populated at λP, likely representing productive and unproductive forms of RPo observed in solution studies. We find that changes in the sequence and length of DNA in the transcription bubble just upstream of the start site (+1) globally alter the network of DNA-RNAP interactions, base stacking, and strand order in the single-stranded DNA of the transcription bubble; these differences propagate beyond the bubble to upstream and downstream DNA. After expanding the transcription bubble by one base (T7A1), the nontemplate strand "scrunches" inside the active site cleft; the template strand bulges outside the cleft at the upstream edge of the bubble. The structures illustrate how limited sequence changes trigger global alterations in the transcription bubble that modulate the RPo lifetime and affect the subsequent steps of the transcription cycle.
History
DepositionApr 23, 2021-
Header (metadata) releaseSep 29, 2021-
Map releaseSep 29, 2021-
UpdateOct 13, 2021-
Current statusOct 13, 2021Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.023
  • Imaged by UCSF Chimera
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  • Surface view colored by radius
  • Surface level: 0.023
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-7mki
  • Surface level: 0.023
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_23895.png

Downloads & links

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Map

FileDownload / File: emd_23895.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Voxel sizeX=Y=Z: 1.03 Å
Density
Contour LevelBy AUTHOR: 0.023 / Movie #1: 0.023
Minimum - Maximum-0.15756056 - 0.23637229
Average (Standard dev.)5.076481e-05 (±0.007736935)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions256256256
Spacing256256256
CellA=B=C: 263.68 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.031.031.03
M x/y/z256256256
origin x/y/z0.0000.0000.000
length x/y/z263.680263.680263.680
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS256256256
D min/max/mean-0.1580.2360.000

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Supplemental data

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Additional map: #1

Fileemd_23895_additional_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #2

Fileemd_23895_half_map_1.map
Projections & Slices
AxesZYX

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Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #1

Fileemd_23895_half_map_2.map
Projections & Slices
AxesZYX

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Slices (1/2)
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Histogram

Histogram (log scale)

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Sample components

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Entire : Escherichia coli RNA polymerase bound to lambda PR promoter DNA (...

EntireName: Escherichia coli RNA polymerase bound to lambda PR promoter DNA (-5G to C)
Components
  • Complex: Escherichia coli RNA polymerase bound to lambda PR promoter DNA (-5G to C)
    • Complex: RNA polymerase
      • Protein or peptide: DNA-directed RNA polymerase subunit alphaPolymerase
      • Protein or peptide: DNA-directed RNA polymerase subunit betaPolymerase
      • Protein or peptide: DNA-directed RNA polymerase subunit beta'Polymerase
      • Protein or peptide: DNA-directed RNA polymerase subunit omegaPolymerase
      • Protein or peptide: RNA polymerase sigma factor RpoD
    • Complex: lambda PR (-5G to C) promoter DNA
      • DNA: Nontemplate strand of lambda PR DNA promoter (-5G to C)
      • DNA: Template strand of lambda PR promoter DNA (-5C to G)
  • Ligand: CHAPSOCHAPS detergent
  • Ligand: MAGNESIUM ION
  • Ligand: ZINC ION

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Supramolecule #1: Escherichia coli RNA polymerase bound to lambda PR promoter DNA (...

SupramoleculeName: Escherichia coli RNA polymerase bound to lambda PR promoter DNA (-5G to C)
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#7

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Supramolecule #2: RNA polymerase

SupramoleculeName: RNA polymerase / type: complex / ID: 2 / Parent: 1 / Macromolecule list: #2-#6
Source (natural)Organism: Escherichia coli (E. coli)
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)

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Supramolecule #3: lambda PR (-5G to C) promoter DNA

SupramoleculeName: lambda PR (-5G to C) promoter DNA / type: complex / ID: 3 / Parent: 1 / Macromolecule list: #1, #7
Source (natural)Organism: Escherichia virus Lambda

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Macromolecule #1: Nontemplate strand of lambda PR DNA promoter (-5G to C)

MacromoleculeName: Nontemplate strand of lambda PR DNA promoter (-5G to C)
type: dna / ID: 1 / Number of copies: 1 / Classification: DNA
Source (natural)Organism: Escherichia virus Lambda
Molecular weightTheoretical: 27.841791 KDa
SequenceString: (DG)(DG)(DA)(DT)(DA)(DA)(DA)(DT)(DA)(DT) (DC)(DT)(DA)(DA)(DC)(DA)(DC)(DC)(DG)(DT) (DG)(DC)(DG)(DT)(DG)(DT)(DT)(DG)(DA) (DC)(DT)(DA)(DT)(DT)(DT)(DT)(DA)(DC)(DC) (DT) (DC)(DT)(DG)(DG)(DC)(DG) ...String:
(DG)(DG)(DA)(DT)(DA)(DA)(DA)(DT)(DA)(DT) (DC)(DT)(DA)(DA)(DC)(DA)(DC)(DC)(DG)(DT) (DG)(DC)(DG)(DT)(DG)(DT)(DT)(DG)(DA) (DC)(DT)(DA)(DT)(DT)(DT)(DT)(DA)(DC)(DC) (DT) (DC)(DT)(DG)(DG)(DC)(DG)(DG)(DT) (DG)(DA)(DT)(DA)(DA)(DT)(DG)(DC)(DT)(DT) (DG)(DC) (DA)(DT)(DG)(DT)(DA)(DC)(DT) (DA)(DA)(DG)(DG)(DA)(DG)(DG)(DT)(DT)(DG) (DT)(DA)(DT) (DG)(DT)(DC)(DG)(DA)(DC) (DC)(DT)(DC)(DG)

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Macromolecule #7: Template strand of lambda PR promoter DNA (-5C to G)

MacromoleculeName: Template strand of lambda PR promoter DNA (-5C to G) / type: dna / ID: 7 / Number of copies: 1 / Classification: DNA
Source (natural)Organism: Escherichia virus Lambda
Molecular weightTheoretical: 27.673764 KDa
SequenceString: (DC)(DG)(DA)(DG)(DG)(DT)(DC)(DG)(DA)(DC) (DA)(DT)(DA)(DC)(DA)(DA)(DC)(DC)(DT)(DC) (DC)(DT)(DT)(DA)(DG)(DT)(DA)(DC)(DA) (DT)(DG)(DC)(DA)(DA)(DG)(DC)(DA)(DT)(DT) (DA) (DT)(DC)(DA)(DC)(DC)(DG) ...String:
(DC)(DG)(DA)(DG)(DG)(DT)(DC)(DG)(DA)(DC) (DA)(DT)(DA)(DC)(DA)(DA)(DC)(DC)(DT)(DC) (DC)(DT)(DT)(DA)(DG)(DT)(DA)(DC)(DA) (DT)(DG)(DC)(DA)(DA)(DG)(DC)(DA)(DT)(DT) (DA) (DT)(DC)(DA)(DC)(DC)(DG)(DC)(DC) (DA)(DG)(DA)(DG)(DG)(DT)(DA)(DA)(DA)(DA) (DT)(DA) (DG)(DT)(DC)(DA)(DA)(DC)(DA) (DC)(DG)(DC)(DA)(DC)(DG)(DG)(DT)(DG)(DT) (DT)(DA)(DG) (DA)(DT)(DA)(DT)(DT)(DT) (DA)(DT)(DC)(DC)

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Macromolecule #2: DNA-directed RNA polymerase subunit alpha

MacromoleculeName: DNA-directed RNA polymerase subunit alpha / type: protein_or_peptide / ID: 2 / Number of copies: 2 / Enantiomer: LEVO / EC number: DNA-directed RNA polymerase
Source (natural)Organism: Escherichia coli (E. coli)
Molecular weightTheoretical: 36.55868 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: MQGSVTEFLK PRLVDIEQVS STHAKVTLEP LERGFGHTLG NALRRILLSS MPGCAVTEVE IDGVLHEYST KEGVQEDILE ILLNLKGLA VRVQGKDEVI LTLNKSGIGP VTAADITHDG DVEIVKPQHV ICHLTDENAS ISMRIKVQRG RGYVPASTRI H SEEDERPI ...String:
MQGSVTEFLK PRLVDIEQVS STHAKVTLEP LERGFGHTLG NALRRILLSS MPGCAVTEVE IDGVLHEYST KEGVQEDILE ILLNLKGLA VRVQGKDEVI LTLNKSGIGP VTAADITHDG DVEIVKPQHV ICHLTDENAS ISMRIKVQRG RGYVPASTRI H SEEDERPI GRLLVDACYS PVERIAYNVE AARVEQRTDL DKLVIEMETN GTIDPEEAIR RAATILAEQL EAFVDLRDVR QP EVKEEKP EFDPILLRPV DDLELTVRSA NCLKAEAIHY IGDLVQRTEV ELLKTPNLGK KSLTEIKDVL ASRGLSLGMR LEN WPPASI ADE

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Macromolecule #3: DNA-directed RNA polymerase subunit beta

MacromoleculeName: DNA-directed RNA polymerase subunit beta / type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO / EC number: DNA-directed RNA polymerase
Source (natural)Organism: Escherichia coli (E. coli)
Molecular weightTheoretical: 150.820875 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: MVYSYTEKKR IRKDFGKRPQ VLDVPYLLSI QLDSFQKFIE QDPEGQYGLE AAFRSVFPIQ SYSGNSELQY VSYRLGEPVF DVQECQIRG VTYSAPLRVK LRLVIYEREA PEGTVKDIKE QEVYMGEIPL MTDNGTFVIN GTERVIVSQL HRSPGVFFDS D KGKTHSSG ...String:
MVYSYTEKKR IRKDFGKRPQ VLDVPYLLSI QLDSFQKFIE QDPEGQYGLE AAFRSVFPIQ SYSGNSELQY VSYRLGEPVF DVQECQIRG VTYSAPLRVK LRLVIYEREA PEGTVKDIKE QEVYMGEIPL MTDNGTFVIN GTERVIVSQL HRSPGVFFDS D KGKTHSSG KVLYNARIIP YRGSWLDFEF DPKDNLFVRI DRRRKLPATI ILRALNYTTE QILDLFFEKV IFEIRDNKLQ ME LVPERLR GETASFDIEA NGKVYVEKGR RITARHIRQL EKDDVKLIEV PVEYIAGKVV AKDYIDESTG ELICAANMEL SLD LLAKLS QSGHKRIETL FTNDLDHGPY ISETLRVDPT NDRLSALVEI YRMMRPGEPP TREAAESLFE NLFFSEDRYD LSAV GRMKF NRSLLREEIE GSGILSKDDI IDVMKKLIDI RNGKGEVDDI DHLGNRRIRS VGEMAENQFR VGLVRVERAV KERLS LGDL DTLMPQDMIN AKPISAAVKE FFGSSQLSQF MDQNNPLSEI THKRRISALG PGGLTRERAG FEVRDVHPTH YGRVCP IET PEGPNIGLIN SLSVYAQTNE YGFLETPYRK VTDGVVTDEI HYLSAIEEGN YVIAQANSNL DEEGHFVEDL VTCRSKG ES SLFSRDQVDY MDVSTQQVVS VGASLIPFLE HDDANRALMG ANMQRQAVPT LRADKPLVGT GMERAVAVDS GVTAVAKR G GVVQYVDASR IVIKVNEDEM YPGEAGIDIY NLTKYTRSNQ NTCINQMPCV SLGEPVERGD VLADGPSTDL GELALGQNM RVAFMPWNGY NFEDSILVSE RVVQEDRFTT IHIQELACVS RDTKLGPEEI TADIPNVGEA ALSKLDESGI VYIGAEVTGG DILVGKVTP KGETQLTPEE KLLRAIFGEK ASDVKDSSLR VPNGVSGTVI DVQVFTRDGV EKDKRALEIE EMQLKQAKKD L SEELQILE AGLFSRIRAV LVAGGVEAEK LDKLPRDRWL ELGLTDEEKQ NQLEQLAEQY DELKHEFEKK LEAKRRKITQ GD DLAPGVL KIVKVYLAVK RRIQPGDKMA GRHGNKGVIS KINPIEDMPY DENGTPVDIV LNPLGVPSRM NIGQILETHL GMA AKGIGD KINAMLKQQQ EVAKLREFIQ RAYDLGADVR QKVDLSTFSD EEVMRLAENL RKGMPIATPV FDGAKEAEIK ELLK LGDLP TSGQIRLYDG RTGEQFERPV TVGYMYMLKL NHLVDDKMHA RSTGSYSLVT QQPLGGKAQF GGQRFGEMEV WALEA YGAA YTLQEMLTVK SDDVNGRTKM YKNIVDGNHQ MEPGMPESFN VLLKEIRSLG INIELEDE

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Macromolecule #4: DNA-directed RNA polymerase subunit beta'

MacromoleculeName: DNA-directed RNA polymerase subunit beta' / type: protein_or_peptide / ID: 4 / Number of copies: 1 / Enantiomer: LEVO / EC number: DNA-directed RNA polymerase
Source (natural)Organism: Escherichia coli (E. coli)
Molecular weightTheoretical: 155.366781 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: MKDLLKFLKA QTKTEEFDAI KIALASPDMI RSWSFGEVKK PETINYRTFK PERDGLFCAR IFGPVKDYEC LCGKYKRLKH RGVICEKCG VEVTQTKVRR ERMGHIELAS PTAHIWFLKS LPSRIGLLLD MPLRDIERVL YFESYVVIEG GMTNLERQQI L TEEQYLDA ...String:
MKDLLKFLKA QTKTEEFDAI KIALASPDMI RSWSFGEVKK PETINYRTFK PERDGLFCAR IFGPVKDYEC LCGKYKRLKH RGVICEKCG VEVTQTKVRR ERMGHIELAS PTAHIWFLKS LPSRIGLLLD MPLRDIERVL YFESYVVIEG GMTNLERQQI L TEEQYLDA LEEFGDEFDA KMGAEAIQAL LKSMDLEQEC EQLREELNET NSETKRKKLT KRIKLLEAFV QSGNKPEWMI LT VLPVLPP DLRPLVPLDG GRFATSDLND LYRRVINRNN RLKRLLDLAA PDIIVRNEKR MLQEAVDALL DNGRRGRAIT GSN KRPLKS LADMIKGKQG RFRQNLLGKR VDYSGRSVIT VGPYLRLHQC GLPKKMALEL FKPFIYGKLE LRGLATTIKA AKKM VEREE AVVWDILDEV IREHPVLLNR APTLHRLGIQ AFEPVLIEGK AIQLHPLVCA AYNADFDGDQ MAVHVPLTLE AQLEA RALM MSTNNILSPA NGEPIIVPSQ DVVLGLYYMT RDCVNAKGEG MVLTGPKEAE RLYRSGLASL HARVKVRITE YEKDAN GEL VAKTSLKDTT VGRAILWMIV PKGLPYSIVN QALGKKAISK MLNTCYRILG LKPTVIFADQ IMYTGFAYAA RSGASVG ID DMVIPEKKHE IISEAEAEVA EIQEQFQSGL VTAGERYNKV IDIWAAANDR VSKAMMDNLQ TETVINRDGQ EEKQVSFN S IYMMADSGAR GSAAQIRQLA GMRGLMAKPD GSIIETPITA NFREGLNVLQ YFISTHGARK GLADTALKTA NSGYLTRRL VDVAQDLVVT EDDCGTHEGI MMTPVIEGGD VKEPLRDRVL GRVTAEDVLK PGTADILVPR NTLLHEQWCD LLEENSVDAV KVRSVVSCD TDFGVCAHCY GRDLARGHII NKGEAIGVIA AQSIGEPGTQ LTMRTFHIGG AASRAAAESS IQVKNKGSIK L SNVKSVVN SSGKLVITSR NTELKLIDEF GRTKESYKVP YGAVLAKGDG EQVAGGETVA NWDPHTMPVI TEVSGFVRFT DM IDGQTIT RQTDELTGLS SLVVLDSAER TAGGKDLRPA LKIVDAQGND VLIPGTDMPA QYFLPGKAIV QLEDGVQISS GDT LARIPQ ESGGTKDITG GLPRVADLFE ARRPKEPAIL AEISGIVSFG KETKGKRRLV ITPVDGSDPY EEMIPKWRQL NVFE GERVE RGDVISDGPE APHDILRLRG VHAVTRYIVN EVQDVYRLQG VKINDKHIEV IVRQMLRKAT IVNAGSSDFL EGEQV EYSR VKIANRELEA NGKVGATYSR DLLGITKASL ATESFISAAS FQETTRVLTE AAVAGKRDEL RGLKENVIVG RLIPAG TGY AYHQDRMRRR AAGEAPAAPQ VTAEDASASL AELLNAGLGG SDNE

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Macromolecule #5: DNA-directed RNA polymerase subunit omega

MacromoleculeName: DNA-directed RNA polymerase subunit omega / type: protein_or_peptide / ID: 5 / Number of copies: 1 / Enantiomer: LEVO / EC number: DNA-directed RNA polymerase
Source (natural)Organism: Escherichia coli (E. coli)
Molecular weightTheoretical: 10.249547 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString:
MARVTVQDAV EKIGNRFDLV LVAARRARQM QVGGKDPLVP EENDKTTVIA LREIEEGLIN NQILDVRERQ EQQEQEAAEL QAVTAIAEG RR

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Macromolecule #6: RNA polymerase sigma factor RpoD

MacromoleculeName: RNA polymerase sigma factor RpoD / type: protein_or_peptide / ID: 6 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Escherichia coli (E. coli)
Molecular weightTheoretical: 70.352242 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: MEQNPQSQLK LLVTRGKEQG YLTYAEVNDH LPEDIVDSDQ IEDIIQMIND MGIQVMEEAP DADDLMLAEN TADEDAAEAA AQVLSSVES EIGRTTDPVR MYMREMGTVE LLTREGEIDI AKRIEDGINQ VQCSVAEYPE AITYLLEQYD RVEAEEARLS D LITGFVDP ...String:
MEQNPQSQLK LLVTRGKEQG YLTYAEVNDH LPEDIVDSDQ IEDIIQMIND MGIQVMEEAP DADDLMLAEN TADEDAAEAA AQVLSSVES EIGRTTDPVR MYMREMGTVE LLTREGEIDI AKRIEDGINQ VQCSVAEYPE AITYLLEQYD RVEAEEARLS D LITGFVDP NAEEDLAPTA THVGSELSQE DLDDDEDEDE EDGDDDSADD DNSIDPELAR EKFAELRAQY VVTRDTIKAK GR SHATAQE EILKLSEVFK QFRLVPKQFD YLVNSMRVMM DRVRTQERLI MKLCVEQCKM PKKNFITLFT GNETSDTWFN AAI AMNKPW SEKLHDVSEE VHRALQKLQQ IEEETGLTIE QVKDINRRMS IGEAKARRAK KEMVEANLRL VISIAKKYTN RGLQ FLDLI QEGNIGLMKA VDKFEYRRGY KFSTYATWWI RQAITRSIAD QARTIRIPVH MIETINKLNR ISRQMLQEMG REPTP EELA ERMLMPEDKI RKVLKIAKEP ISMETPIGDD EDSHLGDFIE DTTLELPLDS ATTESLRAAT HDVLAGLTAR EAKVLR MRF GIDMNTDYTL EEVGKQFDVT RERIRQIEAK ALRKLRHPSR SEVLRSFLDD

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Macromolecule #8: CHAPSO

MacromoleculeName: CHAPSO / type: ligand / ID: 8 / Number of copies: 3 / Formula: 1N7
Molecular weightTheoretical: 631.884 Da
Chemical component information

ChemComp-1N7:
CHAPSO / detergent*YM / CHAPS detergent

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Macromolecule #9: MAGNESIUM ION

MacromoleculeName: MAGNESIUM ION / type: ligand / ID: 9 / Number of copies: 1 / Formula: MG
Molecular weightTheoretical: 24.305 Da

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Macromolecule #10: ZINC ION

MacromoleculeName: ZINC ION / type: ligand / ID: 10 / Number of copies: 2 / Formula: ZN
Molecular weightTheoretical: 65.409 Da

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

BufferpH: 8
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Average electron dose: 75.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.5 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 353304

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