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Yorodumi- EMDB-29683: Cryo-EM structure of 3DVA component 2 of Escherichia coli que-PEC... -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-29683 | |||||||||||||||
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Title | Cryo-EM structure of 3DVA component 2 of Escherichia coli que-PEC (paused elongation complex) RNA Polymerase minus preQ1 ligand | |||||||||||||||
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Keywords | RNA Polymerase / Riboswitch / Transcription / preQ1 | |||||||||||||||
Function / homology | Function and homology information RNA polymerase complex / submerged biofilm formation / cellular response to cell envelope stress / cytosolic DNA-directed RNA polymerase complex / regulation of DNA-templated transcription initiation / bacterial-type flagellum assembly / bacterial-type flagellum-dependent cell motility / nitrate assimilation / transcription elongation factor complex / DNA-directed RNA polymerase complex ...RNA polymerase complex / submerged biofilm formation / cellular response to cell envelope stress / cytosolic DNA-directed RNA polymerase complex / regulation of DNA-templated transcription initiation / bacterial-type flagellum assembly / bacterial-type flagellum-dependent cell motility / nitrate assimilation / transcription elongation factor complex / DNA-directed RNA polymerase complex / regulation of DNA-templated transcription elongation / transcription antitermination / cell motility / DNA-templated transcription initiation / ribonucleoside binding / DNA-directed 5'-3' RNA polymerase activity / DNA-directed RNA polymerase / response to heat / protein-containing complex assembly / intracellular iron ion homeostasis / protein dimerization activity / DNA-templated transcription / magnesium ion binding / DNA binding / zinc ion binding / cytosol / cytoplasm Similarity search - Function | |||||||||||||||
Biological species | Escherichia coli (E. coli) | |||||||||||||||
Method | single particle reconstruction / cryo EM / Resolution: 3.7 Å | |||||||||||||||
Authors | Porta JC / Chauvier A / Deb I / Ellinger E / Frank AT / Meze K / Ohi MD / Walter NG | |||||||||||||||
Funding support | United States, 4 items
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Citation | Journal: Nat Struct Mol Biol / Year: 2023 Title: Structural basis for control of bacterial RNA polymerase pausing by a riboswitch and its ligand. Authors: Adrien Chauvier / Jason C Porta / Indrajit Deb / Emily Ellinger / Katarina Meze / Aaron T Frank / Melanie D Ohi / Nils G Walter / Abstract: Folding of nascent transcripts can be modulated by the RNA polymerase (RNAP) that carries out their transcription, and vice versa. A pause of RNAP during transcription of a preQ riboswitch (termed ...Folding of nascent transcripts can be modulated by the RNA polymerase (RNAP) that carries out their transcription, and vice versa. A pause of RNAP during transcription of a preQ riboswitch (termed que-PEC) is stabilized by a previously characterized template consensus sequence and the ligand-free conformation of the nascent RNA. Ligand binding to the riboswitch induces RNAP pause release and downstream transcription termination; however, the mechanism by which riboswitch folding modulates pausing is unclear. Here, we report single-particle cryo-electron microscopy reconstructions of que-PEC in ligand-free and ligand-bound states. In the absence of preQ, the RNA transcript is in an unexpected hyper-translocated state, preventing downstream nucleotide incorporation. Strikingly, on ligand binding, the riboswitch rotates around its helical axis, expanding the surrounding RNAP exit channel and repositioning the transcript for elongation. Our study reveals the tight coupling by which nascent RNA structures and their ligands can functionally regulate the macromolecular transcription machinery. | |||||||||||||||
History |
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-Structure visualization
Supplemental images |
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-Downloads & links
-EMDB archive
Map data | emd_29683.map.gz | 97.3 MB | EMDB map data format | |
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Header (meta data) | emd-29683-v30.xml emd-29683.xml | 28.7 KB 28.7 KB | Display Display | EMDB header |
FSC (resolution estimation) | emd_29683_fsc.xml | 10.9 KB | Display | FSC data file |
Images | emd_29683.png | 55.3 KB | ||
Others | emd_29683_half_map_1.map.gz emd_29683_half_map_2.map.gz | 95.5 MB 95.5 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-29683 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-29683 | HTTPS FTP |
-Related structure data
Related structure data | 8g2wMC 8f3cC 8g00C 8g1sC 8g4wC 8g7eC 8g8zC C: citing same article (ref.) M: atomic model generated by this map |
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Similar structure data | Similarity search - Function & homologyF&H Search |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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Related items in Molecule of the Month |
-Map
File | Download / File: emd_29683.map.gz / Format: CCP4 / Size: 103 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||
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Voxel size | X=Y=Z: 1 Å | ||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||
Details | EMDB XML:
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-Supplemental data
-Half map: #2
File | emd_29683_half_map_1.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Half map: #1
File | emd_29683_half_map_2.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Sample components
-Entire : Cryo-EM structure of 3DVA component 2 of Escherichia coli que-PEC...
Entire | Name: Cryo-EM structure of 3DVA component 2 of Escherichia coli que-PEC (paused elongation complex) RNA Polymerase minus preQ1 ligand |
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Components |
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-Supramolecule #1: Cryo-EM structure of 3DVA component 2 of Escherichia coli que-PEC...
Supramolecule | Name: Cryo-EM structure of 3DVA component 2 of Escherichia coli que-PEC (paused elongation complex) RNA Polymerase minus preQ1 ligand type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#7 |
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Source (natural) | Organism: Escherichia coli (E. coli) |
Molecular weight | Theoretical: 420 KDa |
-Macromolecule #1: DNA (39-MER)
Macromolecule | Name: DNA (39-MER) / type: dna / ID: 1 / Number of copies: 1 / Classification: DNA |
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Source (natural) | Organism: Escherichia coli (E. coli) |
Molecular weight | Theoretical: 12.063754 KDa |
Sequence | String: (DG)(DG)(DT)(DC)(DA)(DG)(DT)(DA)(DC)(DG) (DT)(DC)(DC)(DA)(DT)(DT)(DA)(DG)(DC)(DT) (DC)(DT)(DT)(DC)(DG)(DG)(DA)(DA)(DG) (DA)(DG)(DA)(DT)(DT)(DC)(DA)(DG)(DA)(DG) |
-Macromolecule #2: DNA (31-MER)
Macromolecule | Name: DNA (31-MER) / type: dna / ID: 2 / Number of copies: 1 / Classification: DNA |
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Source (natural) | Organism: Escherichia coli (E. coli) |
Molecular weight | Theoretical: 9.486079 KDa |
Sequence | String: (DC)(DT)(DC)(DT)(DG)(DA)(DA)(DT)(DC)(DT) (DC)(DT)(DT)(DC)(DC)(DT)(DC)(DG)(DT)(DG) (DT)(DG)(DG)(DT)(DC)(DA)(DG)(DG)(DA) (DC)(DG) |
-Macromolecule #3: DNA-directed RNA polymerase subunit alpha
Macromolecule | Name: DNA-directed RNA polymerase subunit alpha / type: protein_or_peptide / ID: 3 / Number of copies: 2 / Enantiomer: LEVO / EC number: DNA-directed RNA polymerase |
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Source (natural) | Organism: Escherichia coli (E. coli) |
Molecular weight | Theoretical: 25.971531 KDa |
Recombinant expression | Organism: Escherichia coli (E. coli) |
Sequence | String: 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 EAFVDLE UniProtKB: DNA-directed RNA polymerase subunit alpha |
-Macromolecule #4: DNA-directed RNA polymerase subunit beta
Macromolecule | Name: DNA-directed RNA polymerase subunit beta / type: protein_or_peptide / ID: 4 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: Escherichia coli (E. coli) |
Molecular weight | Theoretical: 150.560562 KDa |
Recombinant expression | Organism: Escherichia coli (E. coli) |
Sequence | String: VYSYTEKKRI RKDFGKRPQV LDVPYLLSIQ LDSFQKFIEQ DPEGQYGLEA AFRSVFPIQS YSGNSELQYV SYRLGEPVFD VQECQIRGV TYSAPLRVKL RLVIYEREAP EGTVKDIKEQ EVYMGEIPLM TDNGTFVING TERVIVSQLH RSPGVFFDSD K GKTHSSGK ...String: VYSYTEKKRI RKDFGKRPQV LDVPYLLSIQ LDSFQKFIEQ DPEGQYGLEA AFRSVFPIQS YSGNSELQYV SYRLGEPVFD VQECQIRGV TYSAPLRVKL RLVIYEREAP EGTVKDIKEQ EVYMGEIPLM TDNGTFVING TERVIVSQLH RSPGVFFDSD K GKTHSSGK VLYNARIIPY RGSWLDFEFD PKDNLFVRID RRRKLPATII LRALNYTTEQ ILDLFFEKVI FEIRDNKLQM EL VPERLRG ETASFDIEAN GKVYVEKGRR ITARHIRQLE KDDVKLIEVP VEYIAGKVVA KDYIDESTGE LICAANMELS LDL LAKLSQ SGHKRIETLF TNDLDHGPYI SETLRVDPTN DRLSALVEIY RMMRPGEPPT REAAESLFEN LFFSEDRYDL SAVG RMKFN RSLLREEIEG SGILSKDDII DVMKKLIDIR NGKGEVDDID HLGNRRIRSV GEMAENQFRV GLVRVERAVK ERLSL GDLD TLMPQDMINA KPISAAVKEF FGSSQLSQFM DQNNPLSEIT HKRRISALGP GGLTRERAGF EVRDVHPTHY GRVCPI ETP EGPNIGLINS LSVYAQTNEY GFLETPYRKV TDGVVTDEIH YLSAIEEGNY VIAQANSNLD EEGHFVEDLV TCRSKGE SS LFSRDQVDYM DVSTQQVVSV GASLIPFLEH DDANRALMGA NMQRQAVPTL RADKPLVGTG MERAVAVDSG VTAVAKRG G VVQYVDASRI VIKVNEDEMY PGEAGIDIYN LTKYTRSNQN TCINQMPCVS LGEPVERGDV LADGPSTDLG ELALGQNMR VAFMPWNGYN FEDSILVSER VVQEDRFTTI HIQELACVSR DTKLGPEEIT ADIPNVGEAA LSKLDESGIV YIGAEVTGGD ILVGKVTPK GETQLTPEEK LLRAIFGEKA SDVKDSSLRV PNGVSGTVID VQVFTRDGVE KDKRALEIEE MQLKQAKKDL S EELQILEA GLFSRIRAVL VAGGVEAEKL DKLPRDRWLE LGLTDEEKQN QLEQLAEQYD ELKHEFEKKL EAKRRKITQG DD LAPGVLK IVKVYLAVKR RIQPGDKMAG RHGNKGVISK INPIEDMPYD ENGTPVDIVL NPLGVPSRMN IGQILETHLG MAA KGIGDK INAMLKQQQE VAKLREFIQR AYDLGADVRQ KVDLSTFSDE EVMRLAENLR KGMPIATPVF DGAKEAEIKE LLKL GDLPT SGQIRLYDGR TGEQFERPVT VGYMYMLKLN HLVDDKMHAR STGSYSLVTQ QPLGGKAQFG GQRFGEMEVW ALEAY GAAY TLQEMLTVKS DDVNGRTKMY KNIVDGNHQM EPGMPESFNV LLKEIRSLGI NIELED UniProtKB: DNA-directed RNA polymerase subunit beta |
-Macromolecule #5: DNA-directed RNA polymerase subunit beta'
Macromolecule | Name: DNA-directed RNA polymerase subunit beta' / type: protein_or_peptide / ID: 5 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: Escherichia coli (E. coli) |
Molecular weight | Theoretical: 150.436344 KDa |
Recombinant expression | Organism: Escherichia coli (E. coli) |
Sequence | String: EFDAIKIALA SPDMIRSWSF GEVKKPETIN YRTFKPERDG LFCARIFGPV KDYECLCGKY KRLKHRGVIC EKCGVEVTQT KVRRERMGH IELASPTAHI WFLKSLPSRI GLLLDMPLRD IERVLYFESY VVIEGGMTNL ERQQILTEEQ YLDALEEFGD E FDAKMGAE ...String: EFDAIKIALA SPDMIRSWSF GEVKKPETIN YRTFKPERDG LFCARIFGPV KDYECLCGKY KRLKHRGVIC EKCGVEVTQT KVRRERMGH IELASPTAHI WFLKSLPSRI GLLLDMPLRD IERVLYFESY VVIEGGMTNL ERQQILTEEQ YLDALEEFGD E FDAKMGAE AIQALLKSMD LEQECEQLRE ELNETNSETK RKKLTKRIKL LEAFVQSGNK PEWMILTVLP VLPPDLRPLV PL DGGRFAT SDLNDLYRRV INRNNRLKRL LDLAAPDIIV RNEKRMLQEA VDALLDNGRR GRAITGSNKR PLKSLADMIK GKQ GRFRQN LLGKRVDYSG RSVITVGPYL RLHQCGLPKK MALELFKPFI YGKLELRGLA TTIKAAKKMV EREEAVVWDI LDEV IREHP VLLNRAPTLH RLGIQAFEPV LIEGKAIQLH PLVCAAYNAD FDGDQMAVHV PLTLEAQLEA RALMMSTNNI LSPAN GEPI IVPSQDVVLG LYYMTRDCVN AKGEGMVLTG PKEAERLYRS GLASLHARVK VRITEYEKDA NGELVAKTSL KDTTVG RAI LWMIVPKGLP YSIVNQALGK KAISKMLNTC YRILGLKPTV IFADQIMYTG FAYAARSGAS VGIDDMVIPE KKHEIIS EA EAEVAEIQEQ FQSGLVTAGE RYNKVIDIWA AANDRVSKAM MDNLQTETVI NRDGQEEKQV SFNSIYMMAD SGARGSAA Q IRQLAGMRGL MAKPDGSIIE TPITANFREG LNVLQYFIST HGARKGLADT ALKTANSGYL TRRLVDVAQD LVVTEDDCG THEGIMMTPV IEGGDVKEPL RDRVLGRVTA EDVLKPGTAD ILVPRNTLLH EQWCDLLEEN SVDAVKVRSV VSCDTDFGVC AHCYGRDLA RGHIINKGEA IGVIAAQSIG EPGTQLTMRT FHIGGAASRA AAESSIQVKN KGSIKLSNVK SVVNSSGKLV I TSRNTELK LIDEFGRTKE SYKVPYGAVL AKGDGEQVAG GETVANWDPH TMPVITEVSG FVRFTDMIDG QTITRQTDEL TG LSSLVVL DSAERTAGGK DLRPALKIVD AQGNDVLIPG TDMPAQYFLP GKAIVQLEDG VQISSGDTLA RIPQESGGTK DIT GGLPRV ADLFEARRPK EPAILAEISG IVSFGKETKG KRRLVITPVD GSDPYEEMIP KWRQLNVFEG ERVERGDVIS DGPE APHDI LRLRGVHAVT RYIVNEVQDV YRLQGVKIND KHIEVIVRQM LRKATIVNAG SSDFLEGEQV EYSRVKIANR ELEAN GKVG ATYSRDLLGI TKASLATESF ISAASFQETT RVLTEAAVAG KRDELRGLKE NVIVGRLIPA GTGYAYHQDR MRRR UniProtKB: DNA-directed RNA polymerase subunit beta' |
-Macromolecule #6: DNA-directed RNA polymerase subunit omega
Macromolecule | Name: DNA-directed RNA polymerase subunit omega / type: protein_or_peptide / ID: 6 / Number of copies: 1 / Enantiomer: LEVO / EC number: DNA-directed RNA polymerase |
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Source (natural) | Organism: Escherichia coli (E. coli) |
Molecular weight | Theoretical: 8.963044 KDa |
Recombinant expression | Organism: Escherichia coli (E. coli) |
Sequence | String: ARVTVQDAVE KIGNRFDLVL VAARRARQMQ VGGKDPLVPE ENDKTTVIAL REIEEGLINN QILDVRERQE QQEQEAAEL UniProtKB: DNA-directed RNA polymerase subunit omega |
-Macromolecule #7: RNA (47-MER)
Macromolecule | Name: RNA (47-MER) / type: rna / ID: 7 / Number of copies: 1 / EC number: DNA-directed RNA polymerase |
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Source (natural) | Organism: Escherichia coli (E. coli) |
Molecular weight | Theoretical: 15.059079 KDa |
Sequence | String: GCAGAGGUUC UAGCUACACC CUCUAUAAAA AACUAAGGAC CACACGA |
-Macromolecule #8: MAGNESIUM ION
Macromolecule | Name: MAGNESIUM ION / type: ligand / ID: 8 / Number of copies: 1 / Formula: MG |
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Molecular weight | Theoretical: 24.305 Da |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 5.00 mg/mL | ||||||||||||
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Buffer | pH: 7.5 Component:
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Grid | Model: C-flat-1.2/1.3 / Material: GOLD / Mesh: 400 / Support film - Material: GOLD / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 60 sec. / Pretreatment - Atmosphere: AIR | ||||||||||||
Vitrification | Cryogen name: ETHANE Details: VITRIFICATION WAS CARRIED OUT IN A CHAMBER WITH THE TEMPERATURE SET TO 4 DEGREES CELSIUS.. |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 3.5 µm / Nominal defocus min: 0.5 µm |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN |
Image recording | Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Digitization - Dimensions - Width: 1 pixel / Digitization - Dimensions - Height: 1 pixel / Average electron dose: 62.0 e/Å2 |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
-Image processing
-Atomic model buiding 1
Initial model | PDB ID: Chain - Source name: PDB / Chain - Initial model type: experimental model |
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Refinement | Space: REAL / Protocol: FLEXIBLE FIT / Target criteria: Cross-correlation coefficient |
Output model | PDB-8g2w: |