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- EMDB-66199: Cryo-EM structure of Sigma28-RNAP open promoter complex from Pseu... -

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Basic information

Entry
Database: EMDB / ID: EMD-66199
TitleCryo-EM structure of Sigma28-RNAP open promoter complex from Pseudomonas aeruginosa
Map data
Sample
  • Complex: Sigma28-RNAP open promoter complex
    • Complex: DNA
      • DNA: DNA (54-MER)
      • DNA: DNA (50-MER)
    • Complex: sigma28
      • Protein or peptide: RNA polymerase sigma factor FliA
    • Complex: RNA polymerase
      • Protein or peptide: DNA-directed RNA polymerase subunit omega
      • Protein or peptide: DNA-directed RNA polymerase subunit alpha
      • Protein or peptide: DNA-directed RNA polymerase subunit beta
      • Protein or peptide: DNA-directed RNA polymerase subunit beta'
  • Ligand: MAGNESIUM ION
  • Ligand: ZINC ION
KeywordsP. aeruginosa / Sigma 28 / RNAP / Flagella / fliC / TRANSCRIPTION
Function / homology
Function and homology information


positive regulation of cell motility / sigma factor activity / bacterial-type RNA polymerase core enzyme binding / cytosolic DNA-directed RNA polymerase complex / DNA-directed RNA polymerase complex / DNA-templated transcription initiation / ribonucleoside binding / DNA-directed RNA polymerase / DNA-directed RNA polymerase activity / protein dimerization activity ...positive regulation of cell motility / sigma factor activity / bacterial-type RNA polymerase core enzyme binding / cytosolic DNA-directed RNA polymerase complex / DNA-directed RNA polymerase complex / DNA-templated transcription initiation / ribonucleoside binding / DNA-directed RNA polymerase / DNA-directed RNA polymerase activity / protein dimerization activity / regulation of DNA-templated transcription / DNA-templated transcription / magnesium ion binding / DNA binding / zinc ion binding / cytoplasm
Similarity search - Function
RNA polymerase sigma factor FliA / RNA polymerase sigma factor, FliA/WhiG / Sigma-70 factors family signature 1. / RNA polymerase sigma-70 region 3 / Sigma-70 region 3 / 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 ...RNA polymerase sigma factor FliA / RNA polymerase sigma factor, FliA/WhiG / Sigma-70 factors family signature 1. / RNA polymerase sigma-70 region 3 / Sigma-70 region 3 / 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 beta subunit, bacterial-type / DNA-directed RNA polymerase, alpha subunit / 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, beta subunit, protrusion / RNA polymerase beta subunit / RNA polymerase Rpb1, domain 1 / RNA polymerase Rpb1, domain 1 / 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 Rpb2, domain 2 / RNA polymerase Rpb2, domain 2 / RNA polymerase, N-terminal / RNA polymerase Rpb1, funnel domain superfamily / RNA polymerase I subunit A N-terminus / 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 / DNA-directed RNA polymerase, subunit 2, hybrid-binding domain superfamily / RNA polymerase Rpb2, domain 6
Similarity search - Domain/homology
DNA-directed RNA polymerase subunit alpha / RNA polymerase sigma factor FliA / DNA-directed RNA polymerase subunit beta / DNA-directed RNA polymerase subunit omega / DNA-directed RNA polymerase subunit beta'
Similarity search - Component
Biological speciesPseudomonas aeruginosa (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.4 Å
AuthorsNln S / Kumar V / Sahoo PK / Kandiah E / Jain D
Funding support India, 2 items
OrganizationGrant numberCountry
Science and Engineering Research Board (SERB)SPF/2022/000085 India
Department of Biotechnology (DBT, India)BT/PR36150/INF/22/214/2020 India
CitationJournal: Nucleic Acids Res / Year: 2026
Title: Structural insights into σ28-dependent transcription initiation and its regulation by anti-sigma factor in Pseudomonas aeruginosa.
Authors: Sheenu / Vineet Kumar / Pankaj Kumar Sahoo / Eaazhisai Kandiah / Deepti Jain /
Abstract: Late flagellar genes in Pseudomonas aeruginosa are transcribed by the group 3 sigma factor, FliA (σ28). σ28 drives the expression of flagellin, which assembles into the flagellar filament in this ...Late flagellar genes in Pseudomonas aeruginosa are transcribed by the group 3 sigma factor, FliA (σ28). σ28 drives the expression of flagellin, which assembles into the flagellar filament in this monoflagellated bacterium. This function is suppressed by the anti-sigma factor FlgM. Here, we present the 1.95Å resolution crystal structure of σ28-FlgM complex, along with a 3.4Å structure of σ28RNAP open promoter complex determined using single particle cryo-electron microscopy from P. aeruginosa. The σ28 adopts a compact conformation upon binding to the anti-sigma factor FlgM, which contacts all three domains of the sigma factor. This conformation is neither conducive to interactions with RNA polymerase nor the promoter DNA. The cryo-EM structure reveals base-specific interactions of σ28 domain 4 (σ4) with -35 element, flipping of -11 base of the template strand, novel interactions of template strand with domain 2 (σ2) and 3 (σ3), and partial insertion of sigma finger into the active site cleft, offering unique features of group 3 sigma interactions with promoter DNA. Perturbation of key residues affects transcription in vitro and flagellar phenotypes as well as bacterial motility in vivo. Analysis of the structural data presented here reveals new insights into transcription regulation of late flagellar genes.
History
DepositionSep 13, 2025-
Header (metadata) releaseFeb 25, 2026-
Map releaseFeb 25, 2026-
UpdateFeb 25, 2026-
Current statusFeb 25, 2026Processing site: PDBj / Status: Released

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Structure visualization

Supplemental images

emd_66199.png

Downloads & links

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Map

FileDownload / File: emd_66199.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

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AxesX (Sec.)Y (Row.)Z (Col.)
0.84 Å/pix.
x 256 pix.
= 214.784 Å		0.84 Å/pix.
x 256 pix.
= 214.784 Å		0.84 Å/pix.
x 256 pix.
= 214.784 Å

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Images are generated by Spider.

Voxel sizeX=Y=Z: 0.839 Å
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Histogram (log scale)
Contour LevelBy AUTHOR: 2.7
Minimum - Maximum-8.942128 - 19.616700999999999
Average (Standard dev.)-0.000000000003629 (±1.0)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderZYX
Origin000
Dimensions256256256
Spacing256256256
CellA=B=C: 214.784 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_66199_msk_1.map
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Half map: #2

Fileemd_66199_half_map_1.map
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Half map: #1

Fileemd_66199_half_map_2.map
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Sample components

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Entire : Sigma28-RNAP open promoter complex

EntireName: Sigma28-RNAP open promoter complex
Components
  • Complex: Sigma28-RNAP open promoter complex
    • Complex: DNA
      • DNA: DNA (54-MER)
      • DNA: DNA (50-MER)
    • Complex: sigma28
      • Protein or peptide: RNA polymerase sigma factor FliA
    • Complex: RNA polymerase
      • Protein or peptide: DNA-directed RNA polymerase subunit omega
      • Protein or peptide: DNA-directed RNA polymerase subunit alpha
      • Protein or peptide: DNA-directed RNA polymerase subunit beta
      • Protein or peptide: DNA-directed RNA polymerase subunit beta'
  • Ligand: MAGNESIUM ION
  • Ligand: ZINC ION

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Supramolecule #1: Sigma28-RNAP open promoter complex

SupramoleculeName: Sigma28-RNAP open promoter complex / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#7
Source (natural)Organism: Pseudomonas aeruginosa (bacteria)

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Supramolecule #2: DNA

SupramoleculeName: DNA / type: complex / ID: 2 / Parent: 1 / Macromolecule list: #1-#2

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Supramolecule #3: sigma28

SupramoleculeName: sigma28 / type: complex / ID: 3 / Parent: 1 / Macromolecule list: #4

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Supramolecule #4: RNA polymerase

SupramoleculeName: RNA polymerase / type: complex / ID: 4 / Parent: 1 / Macromolecule list: #3, #5-#7

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Macromolecule #1: DNA (54-MER)

MacromoleculeName: DNA (54-MER) / type: dna / ID: 1 / Number of copies: 1 / Classification: DNA
Source (natural)Organism: Pseudomonas aeruginosa (bacteria)
Molecular weightTheoretical: 16.560641 KDa
SequenceString: (DT)(DT)(DC)(DT)(DC)(DT)(DA)(DA)(DA)(DG) (DC)(DT)(DC)(DC)(DG)(DC)(DC)(DG)(DG)(DG) (DA)(DA)(DA)(DC)(DG)(DC)(DC)(DG)(DA) (DT)(DA)(DA)(DA)(DC)(DA)(DC)(DC)(DA)(DT) (DG) (DA)(DA)(DC)(DG)(DC)(DG) ...String:
(DT)(DT)(DC)(DT)(DC)(DT)(DA)(DA)(DA)(DG) (DC)(DT)(DC)(DC)(DG)(DC)(DC)(DG)(DG)(DG) (DA)(DA)(DA)(DC)(DG)(DC)(DC)(DG)(DA) (DT)(DA)(DA)(DA)(DC)(DA)(DC)(DC)(DA)(DT) (DG) (DA)(DA)(DC)(DG)(DC)(DG)(DA)(DA) (DT)(DT)(DC)(DT)(DT)(DG)

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Macromolecule #2: DNA (50-MER)

MacromoleculeName: DNA (50-MER) / type: dna / ID: 2 / Number of copies: 1 / Classification: DNA
Source (natural)Organism: Pseudomonas aeruginosa (bacteria)
Molecular weightTheoretical: 16.622664 KDa
SequenceString: (DC)(DA)(DA)(DG)(DA)(DA)(DT)(DT)(DC)(DG) (DC)(DG)(DT)(DT)(DC)(DA)(DA)(DC)(DC)(DA) (DC)(DA)(DT)(DT)(DA)(DT)(DC)(DG)(DG) (DC)(DG)(DT)(DT)(DT)(DC)(DC)(DC)(DG)(DG) (DC) (DG)(DG)(DA)(DG)(DC)(DT) ...String:
(DC)(DA)(DA)(DG)(DA)(DA)(DT)(DT)(DC)(DG) (DC)(DG)(DT)(DT)(DC)(DA)(DA)(DC)(DC)(DA) (DC)(DA)(DT)(DT)(DA)(DT)(DC)(DG)(DG) (DC)(DG)(DT)(DT)(DT)(DC)(DC)(DC)(DG)(DG) (DC) (DG)(DG)(DA)(DG)(DC)(DT)(DT)(DT) (DA)(DG)(DA)(DG)(DA)(DA)

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Macromolecule #3: DNA-directed RNA polymerase subunit omega

MacromoleculeName: DNA-directed RNA polymerase subunit omega / type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO / EC number: DNA-directed RNA polymerase
Source (natural)Organism: Pseudomonas aeruginosa (bacteria)
Molecular weightTheoretical: 9.783876 KDa
SequenceString:
MARVTVEDCL DNVDNRFELV MLATKRARQL ATGGKEPKVA WENDKPTVVA LREIASGLVD ENVVQQEDIV EDEPLFAAFD DEANTEAL

UniProtKB: DNA-directed RNA polymerase subunit omega

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Macromolecule #4: RNA polymerase sigma factor FliA

MacromoleculeName: RNA polymerase sigma factor FliA / type: protein_or_peptide / ID: 4 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Pseudomonas aeruginosa (bacteria)
Molecular weightTheoretical: 28.240924 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: GPLHMMTAAS GVRMYSKAQA QNSQEQLIQR YAPLVKRIAY HLLGRLPASV QVEDLMQAGM IGLLEAAKKY DAGKGASFET YAGIRIRGA MLDEVRKGDW APRSVHRNTR MVTDAIRAIE ARTGRDAKDH EVAAELQLSL EDYYGILSDT QGSRLYSFDD L LQDGEHGL ...String:
GPLHMMTAAS GVRMYSKAQA QNSQEQLIQR YAPLVKRIAY HLLGRLPASV QVEDLMQAGM IGLLEAAKKY DAGKGASFET YAGIRIRGA MLDEVRKGDW APRSVHRNTR MVTDAIRAIE ARTGRDAKDH EVAAELQLSL EDYYGILSDT QGSRLYSFDD L LQDGEHGL PEDTSLSHNE PIHGLLDERF QAALADAIAK LPERERLVLA LYYDEELNLK EIGEVLGVSE SRVSQLHSQC AA RLRARLA DWRSAGS

UniProtKB: RNA polymerase sigma factor FliA

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Macromolecule #5: DNA-directed RNA polymerase subunit alpha

MacromoleculeName: DNA-directed RNA polymerase subunit alpha / type: protein_or_peptide / ID: 5 / Number of copies: 2 / Enantiomer: LEVO / EC number: DNA-directed RNA polymerase
Source (natural)Organism: Pseudomonas aeruginosa (bacteria)
Molecular weightTheoretical: 36.694555 KDa
SequenceString: MQSSVNEFLT PRHIDVQVVS QTRAKITLEP LERGFGHTLG NALRRILLSS MPGCAVVEAE IDGVLHEYSA IEGVQEDVIE ILLNLKGLA IKLHGRDEVT LTLAKKGSGV VTAADIQLDH DVEIINGDHV IANLADNGAL NMKLKVARGR GYEPADARQS D EDESRSIG ...String:
MQSSVNEFLT PRHIDVQVVS QTRAKITLEP LERGFGHTLG NALRRILLSS MPGCAVVEAE IDGVLHEYSA IEGVQEDVIE ILLNLKGLA IKLHGRDEVT LTLAKKGSGV VTAADIQLDH DVEIINGDHV IANLADNGAL NMKLKVARGR GYEPADARQS D EDESRSIG RLQLDASFSP VRRVSYVVEN ARVEQRTNLD KLVLDLETNG TLDPEEAIRR AATILQQQLA AFVDLKGDSE PV VEEQEDE IDPILLRPVD DLELTVRSAN CLKAENIYYI GDLIQRTEVE LLKTPNLGKK SLTEIKDVLA SRGLSLGMRL DNW PPASLK KDDKATA

UniProtKB: DNA-directed RNA polymerase subunit alpha

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Macromolecule #6: DNA-directed RNA polymerase subunit beta

MacromoleculeName: DNA-directed RNA polymerase subunit beta / type: protein_or_peptide / ID: 6 / Number of copies: 1 / Enantiomer: LEVO / EC number: DNA-directed RNA polymerase
Source (natural)Organism: Pseudomonas aeruginosa (bacteria)
Molecular weightTheoretical: 151.037047 KDa
SequenceString: MAYSYTEKKR IRKDFSKLPD VMDVPYLLAI QLDSYREFLQ AGATKEQFRD VGLHAAFKSV FPIISYSGNA ALEYVGYRLG EPAFDVKEC VLRGVTFAVP LRVKVRLIIF DRESSNKAIK DIKEQEVYMG EIPLMTENGT FIINGTERVI VSQLHRSPGV F FDHDRGKT ...String:
MAYSYTEKKR IRKDFSKLPD VMDVPYLLAI QLDSYREFLQ AGATKEQFRD VGLHAAFKSV FPIISYSGNA ALEYVGYRLG EPAFDVKEC VLRGVTFAVP LRVKVRLIIF DRESSNKAIK DIKEQEVYMG EIPLMTENGT FIINGTERVI VSQLHRSPGV F FDHDRGKT HSSGKLLYSA RIIPYRGSWL DFEFDPKDCV FVRIDRRRKL PASVLLRALG YSTEEILNAF YATNVFHIKG ET LNLELVP QRLRGEVASI DIKDGSGKVI VEQGRRITAR HINQLEKAGV SQLEVPFDYL IGRTIAKAIV HPATGEIIAE CNT ELTLDL LAKVAKAQVV RIETLYTNDI DCGPFISDTL KIDNTSNQLE ALVEIYRMMR PGEPPTKEAA ETLFGNLFFS AERY DLSAV GRMKFNRRIG RTEIEGPGVL SKEDIIDVLK TLVDIRNGKG IVDDIDHLGN RRVRCVGEMA ENQFRVGLVR VERAV KERL SMAESEGLMP QDLINAKPVA AAIKEFFGSS QLSQFMDQNN PLSEITHKRR VSALGPGGLT RERAGFEVRD VHPTHY GRV CPIETPEGPN IGLINSLATY ARTNKYGFLE SPYRVVKDSL VTDEIVFLSA IEEADHVIAQ ASATLNEKGQ LVDELVA VR HLNEFTVKAP EDVTLMDVSP KQVVSVAASL IPFLEHDDAN RALMGSNMQR QAVPTLRADK PLVGTGMERN VARDSGVC V VARRGGVIDS VDASRVVVRV ADDEVETGEA GVDIYNLTKY TRSNQNTCIN QRPLVSKGDV VARGDILADG PSTDMGELA LGQNMRVAFM PWNGFNFEDS ICLSERVVQE DRFTTIHIQE LTCVARDTKL GPEEITADIP NVGEAALNKL DEAGIVYVGA EVQAGDILV GKVTPKGETQ LTPEEKLLRA IFGEKASDVK DTSLRVPTGT KGTVIDVQVF TRDGVERDSR ALSIEKMQLD Q IRKDLNEE FRIVEGATFE RLRAALVGAK AEGGPALKKG TEITDDYLDG LERGQWFKLR MADDALNEQL EKAQAYISDR RQ LLDDKFE DKKRKLQQGD DLAPGVLKIV KVYLAIKRRI QPGDKMAGRH GNKGVVSVIM PVEDMPHDAN GTPVDIVLNP LGV PSRMNV GQILETHLGL AAKGLGEKIN RMLEEQRKVA ELRKFLHEIY NEIGGREENL DELGDNEILA LAKNLRGGVP MATP VFDGA KEREIKAMLK LADLPESGQM RLFDGRTGNQ FERPTTVGYM YMLKLNHLVD DKMHARSTGS YSLVTQQPLG GKAQF GGQR FGEMEVWALE AYGAAYTLQE MLTVKSDDVN GRTKMYKNIV DGDHRMEAGM PESFNVLIKE IRSLGIDIEL ETE

UniProtKB: DNA-directed RNA polymerase subunit beta

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Macromolecule #7: DNA-directed RNA polymerase subunit beta'

MacromoleculeName: DNA-directed RNA polymerase subunit beta' / type: protein_or_peptide / ID: 7 / Number of copies: 1 / Enantiomer: LEVO / EC number: DNA-directed RNA polymerase
Source (natural)Organism: Pseudomonas aeruginosa (bacteria)
Molecular weightTheoretical: 154.595 KDa
SequenceString: MKDLLNLLKN QGQIEEFDAI RIGLASPEMI RSWSFGEVKK PETINYRTFK PERDGLFCAK IFGPVKDYEC LCGKYKRLKH RGVICEKCG VEVALAKVRR ERMGHIELAS PVAHIWFLKS LPSRIGLLLD MTLRDIERVL YFESYVVIDP GMTTLEKGQL L NDEQYFEA ...String:
MKDLLNLLKN QGQIEEFDAI RIGLASPEMI RSWSFGEVKK PETINYRTFK PERDGLFCAK IFGPVKDYEC LCGKYKRLKH RGVICEKCG VEVALAKVRR ERMGHIELAS PVAHIWFLKS LPSRIGLLLD MTLRDIERVL YFESYVVIDP GMTTLEKGQL L NDEQYFEA LEEFGDDFDA RMGAEAVHEL LNAIDLEHEI GRLREEIPQT NSETKIKKLS KRLKLMEAFQ GSGNKPEWMV LT VLPVLPP DLRPLVPLDG GRFATSDLND LYRRVINRNN RLKRLLDLAA PDIIVRNEKR MLQEAVDALL DNGRRGRAIT GSN KRPLKS LADMIKGKQG RFRQNLLGKR VDYSGRSVIT VGPTLRLHQC GLPKKMALEL FKPFIFGKLE GRGMATTIKA AKKM VEREL PEVWDVLAEV IREHPVLLNR APTLHRLGIQ AFEPVLIEGK AIQLHPLVCA AYNADFDGDQ MAVHVPLTLE AQLEA RALM MSTNNILSPA NGEPIIVPSQ DVVMGLYYMT REAINAKGEG MAFADLQEVD RAYRSGQASL HARVKVRINE KIKGED GQL TANTRIVDTT VGRALLFQVV PAGLPFDVVN QSMKKKAISK LINHCYRVVG LKDTVIFADQ LMYTGFAYST ISGVSIG VN DFVIPDEKAR IINAATDEVK EIESQYASGL VTQGEKYNKV IDLWSKANDE VSKAMMANLS KEKVVDREGK EVDQESFN S MYMMADSGAR GSAAQIRQLA GMRGLMAKPD GSIIETPITA NFREGLNVLQ YFISTHGARK GLADTALKTA NSGYLTRRL VDVAQDLVVT EIDCGTEHGL LMSPHIEGGD VVEPLGERVL GRVIARDVFK PGSDEVIVPA GTLIDEKWVD FLEVMSVDEV VVRSPITCE TRHGICAMCY GRDLARGHRV NIGEAVGVIA AQSIGEPGTQ LTMRTFHIGG AASRTSAADN VQVKNGGTIR L HNLKHVVR ADGALVAVSR SGELAVADDF GRERERYKLP YGAVISVKEG DKVDPGAIVA KWDPHTHPIV TEVDGTVAFV GM EEGITVK RQTDELTGLT NIEVMDPKDR PAAGKDIRPA VKLIDAAGKD LLLPGTDVPA QYFLPANALV NLTDGAKVSI GDV VARIPQ ETSKTRDITG GLPRVADLFE ARRPKEPSIL AEISGTISFG KETKGKRRLV ITPNDGSDPY EELIPKWRHL NVFE GEQVN RGEVISDGPS NPHDILRLLG VSSLAKYIVN EIQDVYRLQG VKINDKHIET ILRQMLRKVE VSESGDSSFI KGDQV ELTQ VLEENEQLGT EDKFPAKYER VLLGITKASL STESFISAAS FQETTRVLTE AAVTGKRDFL RGLKENVVVG RLIPAG TGL AYHSERKRQR DLGKPQRVSA SEAEAALTEA LNSSGN

UniProtKB: DNA-directed RNA polymerase subunit beta'

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Macromolecule #8: MAGNESIUM ION

MacromoleculeName: MAGNESIUM ION / type: ligand / ID: 8 / Number of copies: 1 / Formula: MG
Molecular weightTheoretical: 24.305 Da

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Macromolecule #9: ZINC ION

MacromoleculeName: ZINC ION / type: ligand / ID: 9 / 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 / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeTFS KRIOS
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Average electron dose: 41.34 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.2 µm / Nominal defocus min: 0.8 µm / Nominal magnification: 105000
Sample stageCooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

CTF correctionSoftware - Name: CTFFIND (ver. 4.1.14) / Type: PHASE FLIPPING AND AMPLITUDE CORRECTION
Startup modelType of model: PDB ENTRY
PDB model - PDB ID:

7vf9

Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 3.4 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION / Number images used: 36636
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC
Final 3D classificationSoftware - Name: cryoSPARC
FSC plot (resolution estimation)

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Atomic model buiding 1

RefinementProtocol: RIGID BODY FIT
Output model

PDB-9wsm:
Cryo-EM structure of Sigma28-RNAP open promoter complex from Pseudomonas aeruginosa

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