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Yorodumi- EMDB-13579: Cryo-EM structure of Mycobacterium tuberculosis RNA polymerase ho... -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-13579 | |||||||||
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Title | Cryo-EM structure of Mycobacterium tuberculosis RNA polymerase holoenzyme comprising sigma factor SigB | |||||||||
Map data | M. tuberculosis sigB-RNAP holoenzyme protomer R1 | |||||||||
Sample |
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Function / homology | Function and homology information RNA polymerase core enzyme binding / Antimicrobial action and antimicrobial resistance in Mtb / sigma factor activity / DNA-directed RNA polymerase complex / peptidoglycan-based cell wall / DNA-templated transcription initiation / ribonucleoside binding / DNA-directed 5'-3' RNA polymerase activity / DNA-directed RNA polymerase / response to heat ...RNA polymerase core enzyme binding / Antimicrobial action and antimicrobial resistance in Mtb / sigma factor activity / DNA-directed RNA polymerase complex / peptidoglycan-based cell wall / DNA-templated transcription initiation / ribonucleoside binding / DNA-directed 5'-3' RNA polymerase activity / DNA-directed RNA polymerase / response to heat / protein dimerization activity / response to hypoxia / response to xenobiotic stimulus / response to antibiotic / DNA-templated transcription / positive regulation of DNA-templated transcription / magnesium ion binding / DNA binding / zinc ion binding / plasma membrane / cytoplasm / cytosol Similarity search - Function | |||||||||
Biological species | Mycobacterium tuberculosis H37Rv (bacteria) / Mycobacterium tuberculosis (strain ATCC 25618 / H37Rv) (bacteria) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 3.84 Å | |||||||||
Authors | Brodolin K | |||||||||
Funding support | France, 1 items
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Citation | Journal: Nucleic Acids Res / Year: 2014 Title: Mycobacterium RbpA cooperates with the stress-response σB subunit of RNA polymerase in promoter DNA unwinding. Authors: Yangbo Hu / Zakia Morichaud / Ayyappasamy Sudalaiyadum Perumal / Françoise Roquet-Baneres / Konstantin Brodolin / Abstract: RbpA, a transcriptional activator that is essential for Mycobacterium tuberculosis replication and survival during antibiotic treatment, binds to RNA polymerase (RNAP) in the absence of promoter DNA. ...RbpA, a transcriptional activator that is essential for Mycobacterium tuberculosis replication and survival during antibiotic treatment, binds to RNA polymerase (RNAP) in the absence of promoter DNA. It has been hypothesized that RbpA stimulates housekeeping gene expression by promoting assembly of the σ(A) subunit with core RNAP. Here, using a purified in vitro transcription system of M. tuberculosis, we show that RbpA functions in a promoter-dependent manner as a companion of RNAP essential for promoter DNA unwinding and formation of the catalytically active open promoter complex (RPo). Screening for RbpA activity using a full panel of the M. tuberculosis σ subunits demonstrated that RbpA targets σ(A) and stress-response σ(B), but not the alternative σ subunits from the groups 3 and 4. In contrast to σ(A), the σ(B) subunit activity displayed stringent dependency upon RbpA. These results suggest that RbpA-dependent control of RPo formation provides a mechanism for tuning gene expression during the switch between different physiological states, and in the stress response. #1: Journal: Acta Crystallogr., Sect. D: Biol. Crystallogr. / Year: 2018 Title: Real-space refinement in PHENIX for cryo-EM and crystallography Authors: Afonine PV / Poon BK / Read RJ / Sobolev OV / Terwilliger TC / Urzhumtsev A / Adams PD | |||||||||
History |
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-Structure visualization
Supplemental images |
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-Downloads & links
-EMDB archive
Map data | emd_13579.map.gz | 328 MB | EMDB map data format | |
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Header (meta data) | emd-13579-v30.xml emd-13579.xml | 28.5 KB 28.5 KB | Display Display | EMDB header |
FSC (resolution estimation) | emd_13579_fsc.xml | 15.7 KB | Display | FSC data file |
Images | emd_13579.png | 171.9 KB | ||
Masks | emd_13579_msk_1.map | 347.6 MB | Mask map | |
Others | emd_13579_additional_1.map.gz emd_13579_half_map_1.map.gz emd_13579_half_map_2.map.gz | 167.7 MB 323 MB 323 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-13579 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-13579 | HTTPS FTP |
-Validation report
Summary document | emd_13579_validation.pdf.gz | 987.3 KB | Display | EMDB validaton report |
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Full document | emd_13579_full_validation.pdf.gz | 986.9 KB | Display | |
Data in XML | emd_13579_validation.xml.gz | 23.3 KB | Display | |
Data in CIF | emd_13579_validation.cif.gz | 30.8 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-13579 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-13579 | HTTPS FTP |
-Related structure data
Related structure data | 7pp4MC M: atomic model generated by this map C: citing same article (ref.) |
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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_13579.map.gz / Format: CCP4 / Size: 347.6 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||
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Annotation | M. tuberculosis sigB-RNAP holoenzyme protomer R1 | ||||||||||||||||||||||||||||||||||||
Projections & slices | Image control
Images are generated by Spider. | ||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 1.1 Å | ||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
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-Supplemental data
-Mask #1
File | emd_13579_msk_1.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Additional map: Raw map, M. tuberculosis sigB-RNAP holoenzyme octamer (C1-map)
File | emd_13579_additional_1.map | ||||||||||||
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Annotation | Raw map, M. tuberculosis sigB-RNAP holoenzyme octamer (C1-map) | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: half-map 1
File | emd_13579_half_map_1.map | ||||||||||||
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Annotation | half-map 1 | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: half-map 2
File | emd_13579_half_map_2.map | ||||||||||||
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Annotation | half-map 2 | ||||||||||||
Projections & Slices |
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Density Histograms |
-Sample components
-Entire : RNA polymerase holoenzyme with sigma factor SigB
Entire | Name: RNA polymerase holoenzyme with sigma factor SigB |
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Components |
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-Supramolecule #1: RNA polymerase holoenzyme with sigma factor SigB
Supramolecule | Name: RNA polymerase holoenzyme with sigma factor SigB / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#5 Details: RNA polymerase holoenzyme assembled from individually expressed RNA polymerase core and sigma factor SigB |
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Source (natural) | Organism: Mycobacterium tuberculosis H37Rv (bacteria) |
Recombinant expression | Organism: Escherichia coli (E. coli) / Recombinant strain: BL21 DE3 / Recombinant plasmid: pET28a |
Molecular weight | Theoretical: 400 KDa |
-Macromolecule #1: DNA-directed RNA polymerase subunit alpha
Macromolecule | Name: DNA-directed RNA polymerase subunit alpha / type: protein_or_peptide / ID: 1 / Number of copies: 2 / Enantiomer: LEVO / EC number: DNA-directed RNA polymerase |
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Source (natural) | Organism: Mycobacterium tuberculosis (strain ATCC 25618 / H37Rv) (bacteria) |
Molecular weight | Theoretical: 37.745328 KDa |
Recombinant expression | Organism: Escherichia coli BL21(DE3) (bacteria) |
Sequence | String: MLISQRPTLS EDVLTDNRSQ FVIEPLEPGF GYTLGNSLRR TLLSSIPGAA VTSIRIDGVL HEFTTVPGVK EDVTEIILNL KSLVVSSEE DEPVTMYLRK QGPGEVTAGD IVPPAGVTVH NPGMHIATLN DKGKLEVELV VERGRGYVPA VQNRASGAEI G RIPVDSIY ...String: MLISQRPTLS EDVLTDNRSQ FVIEPLEPGF GYTLGNSLRR TLLSSIPGAA VTSIRIDGVL HEFTTVPGVK EDVTEIILNL KSLVVSSEE DEPVTMYLRK QGPGEVTAGD IVPPAGVTVH NPGMHIATLN DKGKLEVELV VERGRGYVPA VQNRASGAEI G RIPVDSIY SPVLKVTYKV DATRVEQRTD FDKLILDVET KNSISPRDAL ASAGKTLVEL FGLARELNVE AEGIEIGPSP AE ADHIASF ALPIDDLDLT VRSYNCLKRE GVHTVGELVA RTESDLLDIR NFGQKSIDEV KIKLHQLGLS LKDSPPSFDP SEV AGYDVA TGTWSTEGAY DEQDYAETEQ L |
-Macromolecule #2: DNA-directed RNA polymerase subunit beta
Macromolecule | Name: DNA-directed RNA polymerase subunit beta / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO / EC number: DNA-directed RNA polymerase |
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Source (natural) | Organism: Mycobacterium tuberculosis (strain ATCC 25618 / H37Rv) (bacteria) |
Molecular weight | Theoretical: 129.602344 KDa |
Recombinant expression | Organism: Escherichia coli BL21(DE3) (bacteria) |
Sequence | String: MVLADSRQSK TAASPSPSRP QSSSNNSVPG APNRVSFAKL REPLEVPGLL DVQTDSFEWL IGSPRWRESA AERGDVNPVG GLEEVLYEL SPIEDFSGSM SLSFSDPRFD DVKAPVDECK DKDMTYAAPL FVTAEFINNN TGEIKSQTVF MGDFPMMTEK G TFIINGTE ...String: MVLADSRQSK TAASPSPSRP QSSSNNSVPG APNRVSFAKL REPLEVPGLL DVQTDSFEWL IGSPRWRESA AERGDVNPVG GLEEVLYEL SPIEDFSGSM SLSFSDPRFD DVKAPVDECK DKDMTYAAPL FVTAEFINNN TGEIKSQTVF MGDFPMMTEK G TFIINGTE RVVVSQLVRS PGVYFDETID KSTDKTLHSV KVIPSRGAWL EFDVDKRDTV GVRIDRKRRQ PVTVLLKALG WT SEQIVER FGFSEIMRST LEKDNTVGTD EALLDIYRKL RPGEPPTKES AQTLLENLFF KEKRYDLARV GRYKVNKKLG LHV GEPITS STLTEEDVVA TIEYLVRLHE GQTTMTVPGG VEVPVETDDI DHFGNRRLRT VGELIQNQIR VGMSRMERVV RERM TTQDV EAITPQTLIN IRPVVAAIKE FFGTSQLSQF MDQNNPLSGL THKRRLSALG PGGLSRERAG LEVRDVHPSH YGRMC PIET PEGPNIGLIG SLSVYARVNP FGFIETPYRK VVDGVVSDEI VYLTADEEDR HVVAQANSPI DADGRFVEPR VLVRRK AGE VEYVPSSEVD YMDVSPRQMV SVATAMIPFL EHDDANRALM GANMQRQAVP LVRSEAPLVG TGMELRAAID AGDVVVA EE SGVIEEVSAD YITVMHDNGT RRTYRMRKFA RSNHGTCANQ CPIVDAGDRV EAGQVIADGP CTDDGEMALG KNLLVAIM P WEGHNYEDAI ILSNRLVEED VLTSIHIEEH EIDARDTKLG AEEITRDIPN ISDEVLADLD ERGIVRIGAE VRDGDILVG KVTPKGETEL TPEERLLRAI FGEKAREVRD TSLKVPHGES GKVIGIRVFS REDEDELPAG VNELVRVYVA QKRKISDGDK LAGRHGNKG VIGKILPVED MPFLADGTPV DIILNTHGVP RRMNIGQILE THLGWCAHSG WKVDAAKGVP DWAARLPDEL L EAQPNAIV STPVFDGAQE AELQGLLSCT LPNRDGDVLV DADGKAMLFD GRSGEPFPYP VTVGYMYIMK LHHLVDDKIH AR STGPYSM ITQQPLGGKA QFGGQRFGEM ECWAMQAYGA AYTLQELLTI KSDDTVGRVK VYEAIVKGEN IPEPGIPESF KVL LKELQS LCLNVEVLSS DGAAIELREG EDEDLERAAA NLGINLSRNE SASVEDLA |
-Macromolecule #3: DNA-directed RNA polymerase subunit beta'
Macromolecule | Name: DNA-directed RNA polymerase subunit beta' / type: protein_or_peptide / ID: 3 / Details: contains C-terminal 6xHis-tag / Number of copies: 1 / Enantiomer: LEVO / EC number: DNA-directed RNA polymerase |
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Source (natural) | Organism: Mycobacterium tuberculosis (strain ATCC 25618 / H37Rv) (bacteria) |
Molecular weight | Theoretical: 147.797812 KDa |
Recombinant expression | Organism: Escherichia coli BL21(DE3) (bacteria) |
Sequence | String: MLDVNFFDEL RIGLATAEDI RQWSYGEVKK PETINYRTLK PEKDGLFCEK IFGPTRDWEC YCGKYKRVRF KGIICERCGV EVTRAKVRR ERMGHIELAA PVTHIWYFKG VPSRLGYLLD LAPKDLEKII YFAAYVITSV DEEMRHNELS TLEAEMAVER K AVEDQRDG ...String: MLDVNFFDEL RIGLATAEDI RQWSYGEVKK PETINYRTLK PEKDGLFCEK IFGPTRDWEC YCGKYKRVRF KGIICERCGV EVTRAKVRR ERMGHIELAA PVTHIWYFKG VPSRLGYLLD LAPKDLEKII YFAAYVITSV DEEMRHNELS TLEAEMAVER K AVEDQRDG ELEARAQKLE ADLAELEAEG AKADARRKVR DGGEREMRQI RDRAQRELDR LEDIWSTFTK LAPKQLIVDE NL YRELVDR YGEYFTGAMG AESIQKLIEN FDIDAEAESL RDVIRNGKGQ KKLRALKRLK VVAAFQQSGN SPMGMVLDAV PVI PPELRP MVQLDGGRFA TSDLNDLYRR VINRNNRLKR LIDLGAPEII VNNEKRMLQE SVDALFDNGR RGRPVTGPGN RPLK SLSDL LKGKQGRFRQ NLLGKRVDYS GRSVIVVGPQ LKLHQCGLPK LMALELFKPF VMKRLVDLNH AQNIKSAKRM VERQR PQVW DVLEEVIAEH PVLLNRAPTL HRLGIQAFEP MLVEGKAIQL HPLVCEAFNA DFDGDQMAVH LPLSAEAQAE ARILML SSN NILSPASGRP LAMPRLDMVT GLYYLTTEVP GDTGEYQPAS GDHPETGVYS SPAEAIMAAD RGVLSVRAKI KVRLTQL RP PVEIEAELFG HSGWQPGDAW MAETTLGRVM FNELLPLGYP FVNKQMHKKV QAAIINDLAE RYPMIVVAQT VDKLKDAG F YWATRSGVTV SMADVLVPPR KKEILDHYEE RADKVEKQFQ RGALNHDERN EALVEIWKEA TDEVGQALRE HYPDDNPII TIVDSGATGN FTQTRTLAGM KGLVTNPKGE FIPRPVKSSF REGLTVLEYF INTHGARKGL ADTALRTADS GYLTRRLVDV SQDVIVREH DCQTERGIVV ELAERAPDGT LIRDPYIETS AYARTLGTDA VDEAGNVIVE RGQDLGDPEI DALLAAGITQ V KVRSVLTC ATSTGVCATC YGRSMATGKL VDIGEAVGIV AAQSIGEPGT QLTMRTFHQG GVGEDITGGL PRVQELFEAR VP RGKAPIA DVTGRVRLED GERFYKITIV PDDGGEEVVY DKISKRQRLR VFKHEDGSER VLSDGDHVEV GQQLMEGSAD PHE VLRVQG PREVQIHLVR EVQEVYRAQG VSIHDKHIEV IVRQMLRRVT IIDSGSTEFL PGSLIDRAEF EAENRRVVAE GGEP AAGRP VLMGITKASL ATDSWLSAAS FQETTRVLTD AAINCRSDKL NGLKENVIIG KLIPAGTGIN RYRNIAVQPT EEARA AAYT IPSYEDQYYS PDFGAATGAA VPLDDYGYSD YRHHHHHH |
-Macromolecule #4: DNA-directed RNA polymerase subunit omega
Macromolecule | Name: DNA-directed RNA polymerase subunit omega / type: protein_or_peptide / ID: 4 / Number of copies: 1 / Enantiomer: LEVO / EC number: DNA-directed RNA polymerase |
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Source (natural) | Organism: Mycobacterium tuberculosis (strain ATCC 25618 / H37Rv) (bacteria) |
Molecular weight | Theoretical: 11.85114 KDa |
Recombinant expression | Organism: Escherichia coli BL21(DE3) (bacteria) |
Sequence | String: MSISQSDASL AAVPAVDQFD PSSGASGGYD TPLGITNPPI DELLDRVSSK YALVIYAAKR ARQINDYYNQ LGEGILEYVG PLVEPGLQE KPLSIALREI HADLLEHTEG E |
-Macromolecule #5: RNA polymerase sigma factor SigB
Macromolecule | Name: RNA polymerase sigma factor SigB / type: protein_or_peptide / ID: 5 / Details: contains N-terminal 6xHist-tag / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: Mycobacterium tuberculosis (strain ATCC 25618 / H37Rv) (bacteria) |
Molecular weight | Theoretical: 38.572773 KDa |
Recombinant expression | Organism: Escherichia coli BL21(DE3) (bacteria) |
Sequence | String: MGSSHHHHHH SSGLVPRGSH MADAPTRATT SRVDSDLDAQ SPAADLVRVY LNGIGKTALL NAAGEVELAK RIEAGLYAEH LLETRKRLG ENRKRDLAAV VRDGEAARRH LLEANLRLVV SLAKRYTGRG MPLLDLIQEG NLGLIRAMEK FDYTKGFKFS T YATWWIRQ ...String: MGSSHHHHHH SSGLVPRGSH MADAPTRATT SRVDSDLDAQ SPAADLVRVY LNGIGKTALL NAAGEVELAK RIEAGLYAEH LLETRKRLG ENRKRDLAAV VRDGEAARRH LLEANLRLVV SLAKRYTGRG MPLLDLIQEG NLGLIRAMEK FDYTKGFKFS T YATWWIRQ AITRGMADQS RTIRLPVHLV EQVNKLARIK REMHQHLGRE ATDEELAAES GIPIDKINDL LEHSRDPVSL DM PVGSEEE APLGDFIEDA EAMSAENAVI AELLHTDIRS VLATLDEREH QVIRLRFGLD DGQPRTLDQI GKLFGLSRER VRQ IERDVM SKLRHGERAD RLRSYAS |
-Macromolecule #6: ZINC ION
Macromolecule | Name: ZINC ION / type: ligand / ID: 6 / Number of copies: 2 / Formula: ZN |
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Molecular weight | Theoretical: 65.409 Da |
-Macromolecule #7: MAGNESIUM ION
Macromolecule | Name: MAGNESIUM ION / type: ligand / ID: 7 / 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 | 1.4 mg/mL |
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Buffer | pH: 7.9 |
Grid | Model: Quantifoil R2/2 / Pretreatment - Type: GLOW DISCHARGE |
Vitrification | Cryogen name: ETHANE |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Image recording | Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: SUPER-RESOLUTION / Number real images: 3064 / Average electron dose: 49.6 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |