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- EMDB-31403: Cryo-EM structure of Pseudomonas aeruginosa SutA transcription ac... -

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Basic information

Entry
Database: EMDB / ID: EMD-31403
TitleCryo-EM structure of Pseudomonas aeruginosa SutA transcription activation complex
Map data
Sample
  • Complex: Pseudomonas aeruginosa SutA transcription activation complex
    • Complex: Pseudomonas aeruginosa SutA transcription activation
      • 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'
      • Protein or peptide: DNA-directed RNA polymerase subunit omega
      • Protein or peptide: Transcriptional factor SutA
      • Protein or peptide: RNA polymerase sigma factor RpoS
    • Complex: DNA
      • DNA: DNA (70-MER)
      • DNA: DNA (70-MER)
  • Ligand: ZINC ION
  • Ligand: MAGNESIUM ION
Keywordstranscription initiation / Pseudomonas aeruginosa / SutA / transcription activation / sigmaS / RNA polymerase / RNAP beta lobe / TRANSLATION / TRANSCRIPTION-DNA complex
Function / homology
Function and homology information


regulation of response to salt stress / negative regulation of secondary metabolite biosynthetic process / positive regulation of single-species biofilm formation on inanimate substrate / regulation of cellular response to oxidative stress / positive regulation of chemotaxis / regulation of cell motility / sigma factor activity / bacterial-type RNA polymerase core enzyme binding / cytosolic DNA-directed RNA polymerase complex / regulation of cellular response to heat ...regulation of response to salt stress / negative regulation of secondary metabolite biosynthetic process / positive regulation of single-species biofilm formation on inanimate substrate / regulation of cellular response to oxidative stress / positive regulation of chemotaxis / regulation of cell motility / sigma factor activity / bacterial-type RNA polymerase core enzyme binding / cytosolic DNA-directed RNA polymerase complex / regulation of cellular response to heat / DNA-directed RNA polymerase complex / DNA-templated transcription initiation / ribonucleoside binding / DNA-directed RNA polymerase / DNA-directed RNA polymerase activity / protein dimerization activity / DNA-templated transcription / regulation of DNA-templated transcription / magnesium ion binding / DNA binding / zinc ion binding / cytoplasm
Similarity search - Function
: / : / SutA RNAP-binding domain / RNA polymerase sigma factor RpoS / Sigma-70 factors family signature 1. / : / RNA polymerase sigma-70 region 1.2 / Sigma-70 factor, region 1.2 / RNA polymerase sigma-70 region 3 / Sigma-70 region 3 ...: / : / SutA RNAP-binding domain / RNA polymerase sigma factor RpoS / Sigma-70 factors family signature 1. / : / 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 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 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 Rpb2, domain 2 / RNA polymerase Rpb1, funnel domain superfamily / RNA polymerase Rpb2, domain 2 / 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 / Winged helix-like DNA-binding domain superfamily
Similarity search - Domain/homology
DNA-directed RNA polymerase subunit alpha / RNA polymerase sigma factor RpoS / DNA-directed RNA polymerase subunit beta / DNA-directed RNA polymerase subunit omega / Transcriptional regulator SutA RNAP-binding domain-containing protein / DNA-directed RNA polymerase subunit beta'
Similarity search - Component
Biological speciesPseudomonas aeruginosa PAO1 (bacteria) / Pseudomonas aeruginosa (strain ATCC 15692 / DSM 22644 / CIP 104116 / JCM 14847 / LMG 12228 / 1C / PRS 101 / PAO1) (bacteria) / synthetic construct (others)
Methodsingle particle reconstruction / cryo EM / Resolution: 5.8 Å
AuthorsHe DW / You LL
Funding support China, 2 items
OrganizationGrant numberCountry
National Natural Science Foundation of China (NSFC)31670067 China
National Natural Science Foundation of China (NSFC)31822001 China
CitationJournal: Nat Commun / Year: 2022
Title: Pseudomonas aeruginosa SutA wedges RNAP lobe domain open to facilitate promoter DNA unwinding.
Authors: Dingwei He / Linlin You / Xiaoxian Wu / Jing Shi / Aijia Wen / Zhi Yan / Wenhui Mu / Chengli Fang / Yu Feng / Yu Zhang /
Abstract: Pseudomonas aeruginosa (Pae) SutA adapts bacteria to hypoxia and nutrition-limited environment during chronic infection by increasing transcription activity of an RNA polymerase (RNAP) holoenzyme ...Pseudomonas aeruginosa (Pae) SutA adapts bacteria to hypoxia and nutrition-limited environment during chronic infection by increasing transcription activity of an RNA polymerase (RNAP) holoenzyme comprising the stress-responsive σ factor σ (RNAP-σ). SutA shows no homology to previously characterized RNAP-binding proteins. The structure and mode of action of SutA remain unclear. Here we determined cryo-EM structures of Pae RNAP-σ holoenzyme, Pae RNAP-σ holoenzyme complexed with SutA, and Pae RNAP-σ transcription initiation complex comprising SutA. The structures show SutA pinches RNAP-β protrusion and facilitates promoter unwinding by wedging RNAP-β lobe open. Our results demonstrate that SutA clears an energetic barrier to facilitate promoter unwinding of RNAP-σ holoenzyme.
History
DepositionJun 6, 2021-
Header (metadata) releaseJul 27, 2022-
Map releaseJul 27, 2022-
UpdateJun 12, 2024-
Current statusJun 12, 2024Processing site: PDBj / Status: Released

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Structure visualization

Supplemental images

emd_31403.png

Downloads & links

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Map

FileDownload / File: emd_31403.map.gz / Format: CCP4 / Size: 103 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

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AxesZ (Sec.)Y (Row.)X (Col.)
1 Å/pix.
x 300 pix.
= 300. Å		1 Å/pix.
x 300 pix.
= 300. Å		1 Å/pix.
x 300 pix.
= 300. Å

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Images are generated by Spider.

Voxel sizeX=Y=Z: 1 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.0065
Minimum - Maximum-0.016710728 - 0.042323116
Average (Standard dev.)0.00016624092 (±0.001544315)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions300300300
Spacing300300300
CellA=B=C: 300.0 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_31403_msk_1.map
Projections & Slices
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Sample components

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Entire : Pseudomonas aeruginosa SutA transcription activation complex

EntireName: Pseudomonas aeruginosa SutA transcription activation complex
Components
  • Complex: Pseudomonas aeruginosa SutA transcription activation complex
    • Complex: Pseudomonas aeruginosa SutA transcription activation
      • 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'
      • Protein or peptide: DNA-directed RNA polymerase subunit omega
      • Protein or peptide: Transcriptional factor SutA
      • Protein or peptide: RNA polymerase sigma factor RpoS
    • Complex: DNA
      • DNA: DNA (70-MER)
      • DNA: DNA (70-MER)
  • Ligand: ZINC ION
  • Ligand: MAGNESIUM ION

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Supramolecule #1: Pseudomonas aeruginosa SutA transcription activation complex

SupramoleculeName: Pseudomonas aeruginosa SutA transcription activation complex
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#8
Source (natural)Organism: Pseudomonas aeruginosa PAO1 (bacteria)
Molecular weightTheoretical: 433 KDa

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Supramolecule #2: Pseudomonas aeruginosa SutA transcription activation

SupramoleculeName: Pseudomonas aeruginosa SutA transcription activation / type: complex / ID: 2 / Parent: 1 / Macromolecule list: #1-#6

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Supramolecule #3: DNA

SupramoleculeName: DNA / type: complex / ID: 3 / Parent: 1 / Macromolecule list: #7-#8

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Macromolecule #1: DNA-directed RNA polymerase subunit alpha

MacromoleculeName: DNA-directed RNA polymerase subunit alpha / type: protein_or_peptide / ID: 1 / Number of copies: 2 / Enantiomer: LEVO / EC number: DNA-directed RNA polymerase
Source (natural)Organism: Pseudomonas aeruginosa (strain ATCC 15692 / DSM 22644 / CIP 104116 / JCM 14847 / LMG 12228 / 1C / PRS 101 / PAO1) (bacteria)
Strain: ATCC 15692 / DSM 22644 / CIP 104116 / JCM 14847 / LMG 12228 / 1C / PRS 101 / PAO1
Molecular weightTheoretical: 38.264258 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: MGHHHHHHHH HHMQSSVNEF LTPRHIDVQV VSQTRAKITL EPLERGFGHT LGNALRRILL SSMPGCAVVE AEIDGVLHEY SAIEGVQED VIEILLNLKG LAIKLHGRDE VTLTLAKKGS GVVTAADIQL DHDVEIINGD HVIANLADNG ALNMKLKVAR G RGYEPADA ...String:
MGHHHHHHHH HHMQSSVNEF LTPRHIDVQV VSQTRAKITL EPLERGFGHT LGNALRRILL SSMPGCAVVE AEIDGVLHEY SAIEGVQED VIEILLNLKG LAIKLHGRDE VTLTLAKKGS GVVTAADIQL DHDVEIINGD HVIANLADNG ALNMKLKVAR G RGYEPADA RQSDEDESRS IGRLQLDASF SPVRRVSYVV ENARVEQRTN LDKLVLDLET NGTLDPEEAI RRAATILQQQ LA AFVDLKG DSEPVVEEQE DEIDPILLRP VDDLELTVRS ANCLKAENIY YIGDLIQRTE VELLKTPNLG KKSLTEIKDV LAS RGLSLG MRLDNWPPAS LKKDDKATA

UniProtKB: DNA-directed RNA polymerase subunit alpha

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Macromolecule #2: DNA-directed RNA polymerase subunit beta

MacromoleculeName: 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: Pseudomonas aeruginosa (strain ATCC 15692 / DSM 22644 / CIP 104116 / JCM 14847 / LMG 12228 / 1C / PRS 101 / PAO1) (bacteria)
Strain: ATCC 15692 / DSM 22644 / CIP 104116 / JCM 14847 / LMG 12228 / 1C / PRS 101 / PAO1
Molecular weightTheoretical: 151.225297 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: MGMAYSYTEK KRIRKDFSKL PDVMDVPYLL AIQLDSYREF LQAGATKEQF RDVGLHAAFK SVFPIISYSG NAALEYVGYR LGEPAFDVK ECVLRGVTFA VPLRVKVRLI IFDRESSNKA IKDIKEQEVY MGEIPLMTEN GTFIINGTER VIVSQLHRSP G VFFDHDRG ...String:
MGMAYSYTEK KRIRKDFSKL PDVMDVPYLL AIQLDSYREF LQAGATKEQF RDVGLHAAFK SVFPIISYSG NAALEYVGYR LGEPAFDVK ECVLRGVTFA VPLRVKVRLI IFDRESSNKA IKDIKEQEVY MGEIPLMTEN GTFIINGTER VIVSQLHRSP G VFFDHDRG KTHSSGKLLY SARIIPYRGS WLDFEFDPKD CVFVRIDRRR KLPASVLLRA LGYSTEEILN AFYATNVFHI KG ETLNLEL VPQRLRGEVA SIDIKDGSGK VIVEQGRRIT ARHINQLEKA GVSQLEVPFD YLIGRTIAKA IVHPATGEII AEC NTELTL DLLAKVAKAQ VVRIETLYTN DIDCGPFISD TLKIDNTSNQ LEALVEIYRM MRPGEPPTKE AAETLFGNLF FSAE RYDLS AVGRMKFNRR IGRTEIEGPG VLSKEDIIDV LKTLVDIRNG KGIVDDIDHL GNRRVRCVGE MAENQFRVGL VRVER AVKE RLSMAESEGL MPQDLINAKP VAAAIKEFFG SSQLSQFMDQ NNPLSEITHK RRVSALGPGG LTRERAGFEV RDVHPT HYG RVCPIETPEG PNIGLINSLA TYARTNKYGF LESPYRVVKD SLVTDEIVFL SAIEEADHVI AQASATLNEK GQLVDEL VA VRHLNEFTVK APEDVTLMDV SPKQVVSVAA SLIPFLEHDD ANRALMGSNM QRQAVPTLRA DKPLVGTGME RNVARDSG V CVVARRGGVI DSVDASRVVV RVADDEVETG EAGVDIYNLT KYTRSNQNTC INQRPLVSKG DVVARGDILA DGPSTDMGE LALGQNMRVA FMPWNGFNFE DSICLSERVV QEDRFTTIHI QELTCVARDT KLGPEEITAD IPNVGEAALN KLDEAGIVYV GAEVQAGDI LVGKVTPKGE TQLTPEEKLL RAIFGEKASD VKDTSLRVPT GTKGTVIDVQ VFTRDGVERD SRALSIEKMQ L DQIRKDLN EEFRIVEGAT FERLRAALVG AKAEGGPALK KGTEITDDYL DGLERGQWFK LRMADDALNE QLEKAQAYIS DR RQLLDDK FEDKKRKLQQ GDDLAPGVLK IVKVYLAIKR RIQPGDKMAG RHGNKGVVSV IMPVEDMPHD ANGTPVDIVL NPL GVPSRM NVGQILETHL GLAAKGLGEK INRMLEEQRK VAELRKFLHE IYNEIGGREE NLDELGDNEI LALAKNLRGG VPMA TPVFD GAKEREIKAM LKLADLPESG QMRLFDGRTG NQFERPTTVG YMYMLKLNHL VDDKMHARST GSYSLVTQQP LGGKA QFGG QRFGEMEVWA LEAYGAAYTL QEMLTVKSDD VNGRTKMYKN IVDGDHRMEA GMPESFNVLI KEIRSLGIDI ELETE

UniProtKB: DNA-directed RNA polymerase subunit beta

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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: Pseudomonas aeruginosa (strain ATCC 15692 / DSM 22644 / CIP 104116 / JCM 14847 / LMG 12228 / 1C / PRS 101 / PAO1) (bacteria)
Strain: ATCC 15692 / DSM 22644 / CIP 104116 / JCM 14847 / LMG 12228 / 1C / PRS 101 / PAO1
Molecular weightTheoretical: 156.038516 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: MLKDLLNLLK NQGQIEEFDA IRIGLASPEM IRSWSFGEVK KPETINYRTF KPERDGLFCA KIFGPVKDYE CLCGKYKRLK HRGVICEKC GVEVALAKVR RERMGHIELA SPVAHIWFLK SLPSRIGLLL DMTLRDIERV LYFESYVVID PGMTTLEKGQ L LNDEQYFE ...String:
MLKDLLNLLK NQGQIEEFDA IRIGLASPEM IRSWSFGEVK KPETINYRTF KPERDGLFCA KIFGPVKDYE CLCGKYKRLK HRGVICEKC GVEVALAKVR RERMGHIELA SPVAHIWFLK SLPSRIGLLL DMTLRDIERV LYFESYVVID PGMTTLEKGQ L LNDEQYFE ALEEFGDDFD ARMGAEAVHE LLNAIDLEHE IGRLREEIPQ TNSETKIKKL SKRLKLMEAF QGSGNKPEWM VL TVLPVLP PDLRPLVPLD GGRFATSDLN DLYRRVINRN NRLKRLLDLA APDIIVRNEK RMLQEAVDAL LDNGRRGRAI TGS NKRPLK SLADMIKGKQ GRFRQNLLGK RVDYSGRSVI TVGPTLRLHQ CGLPKKMALE LFKPFIFGKL EGRGMATTIK AAKK MVERE LPEVWDVLAE VIREHPVLLN RAPTLHRLGI QAFEPVLIEG KAIQLHPLVC AAYNADFDGD QMAVHVPLTL EAQLE ARAL MMSTNNILSP ANGEPIIVPS QDVVMGLYYM TREAINAKGE GMAFADLQEV DRAYRSGQAS LHARVKVRIN EKIKGE DGQ LTANTRIVDT TVGRALLFQV VPAGLPFDVV NQSMKKKAIS KLINHCYRVV GLKDTVIFAD QLMYTGFAYS TISGVSI GV NDFVIPDEKA RIINAATDEV KEIESQYASG LVTQGEKYNK VIDLWSKAND EVSKAMMANL SKEKVVDREG KEVDQESF N SMYMMADSGA RGSAAQIRQL AGMRGLMAKP DGSIIETPIT ANFREGLNVL QYFISTHGAR KGLADTALKT ANSGYLTRR LVDVAQDLVV TEIDCGTEHG LLMSPHIEGG DVVEPLGERV LGRVIARDVF KPGSDEVIVP AGTLIDEKWV DFLEVMSVDE VVVRSPITC ETRHGICAMC YGRDLARGHR VNIGEAVGVI AAQSIGEPGT QLTMRTFHIG GAASRTSAAD NVQVKNGGTI R LHNLKHVV RADGALVAVS RSGELAVADD FGRERERYKL PYGAVISVKE GDKVDPGAIV AKWDPHTHPI VTEVDGTVAF VG MEEGITV KRQTDELTGL TNIEVMDPKD RPAAGKDIRP AVKLIDAAGK DLLLPGTDVP AQYFLPANAL VNLTDGAKVS IGD VVARIP QETSKTRDIT GGLPRVADLF EARRPKEPSI LAEISGTISF GKETKGKRRL VITPNDGSDP YEELIPKWRH LNVF EGEQV NRGEVISDGP SNPHDILRLL GVSSLAKYIV NEIQDVYRLQ GVKINDKHIE TILRQMLRKV EVSESGDSSF IKGDQ VELT QVLEENEQLG TEDKFPAKYE RVLLGITKAS LSTESFISAA SFQETTRVLT EAAVTGKRDF LRGLKENVVV GRLIPA GTG LAYHSERKRQ RDLGKPQRVS ASEAEAALTE ALNSSGNGSG SWSHPQFEK

UniProtKB: DNA-directed RNA polymerase subunit beta'

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Macromolecule #4: DNA-directed RNA polymerase subunit omega

MacromoleculeName: DNA-directed RNA polymerase subunit omega / type: protein_or_peptide / ID: 4 / Number of copies: 1 / Enantiomer: LEVO / EC number: DNA-directed RNA polymerase
Source (natural)Organism: Pseudomonas aeruginosa (strain ATCC 15692 / DSM 22644 / CIP 104116 / JCM 14847 / LMG 12228 / 1C / PRS 101 / PAO1) (bacteria)
Strain: ATCC 15692 / DSM 22644 / CIP 104116 / JCM 14847 / LMG 12228 / 1C / PRS 101 / PAO1
Molecular weightTheoretical: 9.783876 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString:
MARVTVEDCL DNVDNRFELV MLATKRARQL ATGGKEPKVA WENDKPTVVA LREIASGLVD ENVVQQEDIV EDEPLFAAFD DEANTEAL

UniProtKB: DNA-directed RNA polymerase subunit omega

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Macromolecule #5: Transcriptional factor SutA

MacromoleculeName: Transcriptional factor SutA / type: protein_or_peptide / ID: 5 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Pseudomonas aeruginosa (strain ATCC 15692 / DSM 22644 / CIP 104116 / JCM 14847 / LMG 12228 / 1C / PRS 101 / PAO1) (bacteria)
Strain: ATCC 15692 / DSM 22644 / CIP 104116 / JCM 14847 / LMG 12228 / 1C / PRS 101 / PAO1
Molecular weightTheoretical: 11.497365 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString:
GAMGMSEEEL EQDELDGADE DDGEELAAAD DGEADSGDGD EAPAPGKKAK AAVVEEELPS VEAKQKERDA LAKAMEEFLS RGGKVQEIE PNVVADPPKK PDSKYGSRPI

UniProtKB: Transcriptional regulator SutA RNAP-binding domain-containing protein

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Macromolecule #6: RNA polymerase sigma factor RpoS

MacromoleculeName: RNA polymerase sigma factor RpoS / type: protein_or_peptide / ID: 6 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Pseudomonas aeruginosa (strain ATCC 15692 / DSM 22644 / CIP 104116 / JCM 14847 / LMG 12228 / 1C / PRS 101 / PAO1) (bacteria)
Strain: ATCC 15692 / DSM 22644 / CIP 104116 / JCM 14847 / LMG 12228 / 1C / PRS 101 / PAO1
Molecular weightTheoretical: 39.361504 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: MALKKEGPEF DHDDEVLLLE PGIMLDESSA DEQPSPRATP KATTSFSSKQ HKHIDYTRAL DATQLYLNEI GFSPLLTPEE EVHFARLAQ KGDPAGRKRM IESNLRLVVK IARRYVNRGL SLLDLIEEGN LGLIRAVEKF DPERGFRFST YATWWIRQTI E RAIMNQTR ...String:
MALKKEGPEF DHDDEVLLLE PGIMLDESSA DEQPSPRATP KATTSFSSKQ HKHIDYTRAL DATQLYLNEI GFSPLLTPEE EVHFARLAQ KGDPAGRKRM IESNLRLVVK IARRYVNRGL SLLDLIEEGN LGLIRAVEKF DPERGFRFST YATWWIRQTI E RAIMNQTR TIRLPIHVVK ELNVYLRAAR ELTHKLDHEP SPEEIANLLE KPVAEVKRML GLNERVTSVD VSLGPDSDKT LL DTLTDDR PTDPCELLQD DDLSESIDQW LTELTDKQRE VVIRRFGLRG HESSTLEEVG QEIGLTRERV RQIQVEALKR LRE ILEKNG LSSDALFQLE HHHHHH

UniProtKB: RNA polymerase sigma factor RpoS

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Macromolecule #7: DNA (70-MER)

MacromoleculeName: DNA (70-MER) / type: dna / ID: 7 / Number of copies: 1 / Classification: DNA
Source (natural)Organism: synthetic construct (others)
Molecular weightTheoretical: 21.514742 KDa
SequenceString: (DG)(DC)(DT)(DT)(DT)(DC)(DG)(DT)(DC)(DT) (DC)(DA)(DA)(DC)(DC)(DG)(DA)(DG)(DA)(DG) (DG)(DC)(DC)(DG)(DC)(DG)(DC)(DA)(DT) (DT)(DC)(DT)(DA)(DC)(DA)(DG)(DC)(DA)(DT) (DC) (DG)(DT)(DG)(DT)(DG)(DG) ...String:
(DG)(DC)(DT)(DT)(DT)(DC)(DG)(DT)(DC)(DT) (DC)(DA)(DA)(DC)(DC)(DG)(DA)(DG)(DA)(DG) (DG)(DC)(DC)(DG)(DC)(DG)(DC)(DA)(DT) (DT)(DC)(DT)(DA)(DC)(DA)(DG)(DC)(DA)(DT) (DC) (DG)(DT)(DG)(DT)(DG)(DG)(DT)(DT) (DC)(DC)(DG)(DT)(DC)(DA)(DA)(DG)(DC)(DG) (DT)(DT) (DA)(DT)(DT)(DT)(DT)(DC)(DG) (DA)(DA)(DA)

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Macromolecule #8: DNA (70-MER)

MacromoleculeName: DNA (70-MER) / type: dna / ID: 8 / Number of copies: 1 / Classification: DNA
Source (natural)Organism: synthetic construct (others)
Molecular weightTheoretical: 21.648875 KDa
SequenceString: (DT)(DT)(DT)(DC)(DG)(DA)(DA)(DA)(DA)(DT) (DA)(DA)(DC)(DG)(DC)(DT)(DT)(DG)(DA)(DC) (DG)(DG)(DA)(DA)(DC)(DC)(DA)(DC)(DA) (DC)(DG)(DA)(DT)(DG)(DC)(DT)(DG)(DT)(DA) (DG) (DA)(DA)(DT)(DG)(DC)(DG) ...String:
(DT)(DT)(DT)(DC)(DG)(DA)(DA)(DA)(DA)(DT) (DA)(DA)(DC)(DG)(DC)(DT)(DT)(DG)(DA)(DC) (DG)(DG)(DA)(DA)(DC)(DC)(DA)(DC)(DA) (DC)(DG)(DA)(DT)(DG)(DC)(DT)(DG)(DT)(DA) (DG) (DA)(DA)(DT)(DG)(DC)(DG)(DC)(DG) (DG)(DC)(DC)(DT)(DC)(DT)(DC)(DG)(DG)(DT) (DT)(DG) (DA)(DG)(DA)(DC)(DG)(DA)(DA) (DA)(DG)(DC)

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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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Macromolecule #10: MAGNESIUM ION

MacromoleculeName: MAGNESIUM ION / type: ligand / ID: 10 / Number of copies: 1 / Formula: MG
Molecular weightTheoretical: 24.305 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

Concentration13 mg/mL
BufferpH: 7.5
GridModel: C-flat-1.2/1.3 / Material: COPPER / Mesh: 400 / Support film - Material: CARBON / Support film - topology: HOLEY / Support film - Film thickness: 4 / Pretreatment - Type: PLASMA CLEANING / Pretreatment - Time: 120 sec. / Pretreatment - Atmosphere: OTHER
VitrificationCryogen name: ETHANE / Chamber humidity: 95 % / Chamber temperature: 283 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: SUPER-RESOLUTION / Average electron dose: 60.8 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 0.0 mm / Nominal defocus max: 2.2 µm / Nominal defocus min: 1.2 µm / Nominal magnification: 22500
Sample stageCooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: PDB ENTRY
PDB model - PDB ID:

7khb

Final reconstructionResolution.type: BY AUTHOR / Resolution: 5.8 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.0) / Number images used: 112959
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.0)
Final angle assignmentType: ANGULAR RECONSTITUTION / Software - Name: RELION (ver. 3.0)
Final 3D classificationNumber classes: 4 / Software - Name: RELION (ver. 3.0)
FSC plot (resolution estimation)

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