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Yorodumi- EMDB-11679: Structure of human mitochondrial RNA polymerase in complex with I... -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-11679 | ||||||||||||||||||||||||||||||||||||||||||||||||
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Title | Structure of human mitochondrial RNA polymerase in complex with IMT inhibitor. | ||||||||||||||||||||||||||||||||||||||||||||||||
Map data | Post-processed main map. | ||||||||||||||||||||||||||||||||||||||||||||||||
Sample |
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Keywords | Mitochondria / Transcription / Polymerase / Inhibitor | ||||||||||||||||||||||||||||||||||||||||||||||||
Function / homology | Function and homology information Mitochondrial transcription initiation / mitochondrial DNA-directed RNA polymerase complex / mitochondrial promoter sequence-specific DNA binding / transcription initiation at mitochondrial promoter / mitochondrial transcription / DNA primase activity / mitochondrial nucleoid / Transcriptional activation of mitochondrial biogenesis / DNA-directed 5'-3' RNA polymerase activity / DNA-directed RNA polymerase ...Mitochondrial transcription initiation / mitochondrial DNA-directed RNA polymerase complex / mitochondrial promoter sequence-specific DNA binding / transcription initiation at mitochondrial promoter / mitochondrial transcription / DNA primase activity / mitochondrial nucleoid / Transcriptional activation of mitochondrial biogenesis / DNA-directed 5'-3' RNA polymerase activity / DNA-directed RNA polymerase / 3'-5'-RNA exonuclease activity / sequence-specific DNA binding / mitochondrial matrix / protein-containing complex / mitochondrion / RNA binding Similarity search - Function | ||||||||||||||||||||||||||||||||||||||||||||||||
Biological species | Homo sapiens (human) | ||||||||||||||||||||||||||||||||||||||||||||||||
Method | single particle reconstruction / cryo EM / Resolution: 3.5 Å | ||||||||||||||||||||||||||||||||||||||||||||||||
Authors | Hillen HS / Bonekamp N | ||||||||||||||||||||||||||||||||||||||||||||||||
Funding support | Sweden, Germany, 15 items
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Citation | Journal: Nature / Year: 2020 Title: Small-molecule inhibitors of human mitochondrial DNA transcription. Authors: Nina A Bonekamp / Bradley Peter / Hauke S Hillen / Andrea Felser / Tim Bergbrede / Axel Choidas / Moritz Horn / Anke Unger / Raffaella Di Lucrezia / Ilian Atanassov / Xinping Li / Uwe Koch / ...Authors: Nina A Bonekamp / Bradley Peter / Hauke S Hillen / Andrea Felser / Tim Bergbrede / Axel Choidas / Moritz Horn / Anke Unger / Raffaella Di Lucrezia / Ilian Atanassov / Xinping Li / Uwe Koch / Sascha Menninger / Joanna Boros / Peter Habenberger / Patrick Giavalisco / Patrick Cramer / Martin S Denzel / Peter Nussbaumer / Bert Klebl / Maria Falkenberg / Claes M Gustafsson / Nils-Göran Larsson / Abstract: Altered expression of mitochondrial DNA (mtDNA) occurs in ageing and a range of human pathologies (for example, inborn errors of metabolism, neurodegeneration and cancer). Here we describe first-in- ...Altered expression of mitochondrial DNA (mtDNA) occurs in ageing and a range of human pathologies (for example, inborn errors of metabolism, neurodegeneration and cancer). Here we describe first-in-class specific inhibitors of mitochondrial transcription (IMTs) that target the human mitochondrial RNA polymerase (POLRMT), which is essential for biogenesis of the oxidative phosphorylation (OXPHOS) system. The IMTs efficiently impair mtDNA transcription in a reconstituted recombinant system and cause a dose-dependent inhibition of mtDNA expression and OXPHOS in cell lines. To verify the cellular target, we performed exome sequencing of mutagenized cells and identified a cluster of amino acid substitutions in POLRMT that cause resistance to IMTs. We obtained a cryo-electron microscopy (cryo-EM) structure of POLRMT bound to an IMT, which further defined the allosteric binding site near the active centre cleft of POLRMT. The growth of cancer cells and the persistence of therapy-resistant cancer stem cells has previously been reported to depend on OXPHOS, and we therefore investigated whether IMTs have anti-tumour effects. Four weeks of oral treatment with an IMT is well-tolerated in mice and does not cause OXPHOS dysfunction or toxicity in normal tissues, despite inducing a strong anti-tumour response in xenografts of human cancer cells. In summary, IMTs provide a potent and specific chemical biology tool to study the role of mtDNA expression in physiology and disease. | ||||||||||||||||||||||||||||||||||||||||||||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | EM map: SurfViewMolmilJmol/JSmol |
Supplemental images |
-Downloads & links
-EMDB archive
Map data | emd_11679.map.gz | 4.4 MB | EMDB map data format | |
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Header (meta data) | emd-11679-v30.xml emd-11679.xml | 22.7 KB 22.7 KB | Display Display | EMDB header |
FSC (resolution estimation) | emd_11679_fsc.xml | 8.6 KB | Display | FSC data file |
Images | emd_11679.png | 55.3 KB | ||
Masks | emd_11679_msk_1.map | 52.7 MB | Mask map | |
Filedesc metadata | emd-11679.cif.gz | 7.1 KB | ||
Others | emd_11679_half_map_1.map.gz emd_11679_half_map_2.map.gz | 40.8 MB 40.8 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-11679 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-11679 | HTTPS FTP |
-Validation report
Summary document | emd_11679_validation.pdf.gz | 762.7 KB | Display | EMDB validaton report |
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Full document | emd_11679_full_validation.pdf.gz | 762.2 KB | Display | |
Data in XML | emd_11679_validation.xml.gz | 13.8 KB | Display | |
Data in CIF | emd_11679_validation.cif.gz | 18.6 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-11679 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-11679 | HTTPS FTP |
-Related structure data
Related structure data | 7a8pMC M: atomic model generated by this map C: citing same article (ref.) |
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Similar structure data |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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Related items in Molecule of the Month |
-Map
File | Download / File: emd_11679.map.gz / Format: CCP4 / Size: 52.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | Post-processed main map. | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Projections & slices | Image control
Images are generated by Spider. | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 0.834 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
-Mask #1
File | emd_11679_msk_1.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Half map: Refinement half-map 1.
File | emd_11679_half_map_1.map | ||||||||||||
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Annotation | Refinement half-map 1. | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: Refinement half-map 2.
File | emd_11679_half_map_2.map | ||||||||||||
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Annotation | Refinement half-map 2. | ||||||||||||
Projections & Slices |
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Density Histograms |
-Sample components
-Entire : Human POLRMT (lacking residues 1-104) in complex with IMT inhibitor.
Entire | Name: Human POLRMT (lacking residues 1-104) in complex with IMT inhibitor. |
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Components |
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-Supramolecule #1: Human POLRMT (lacking residues 1-104) in complex with IMT inhibitor.
Supramolecule | Name: Human POLRMT (lacking residues 1-104) in complex with IMT inhibitor. type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1 |
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Source (natural) | Organism: Homo sapiens (human) / Organelle: Mitochondria |
Molecular weight | Theoretical: 128 KDa |
-Macromolecule #1: DNA-directed RNA polymerase, mitochondrial
Macromolecule | Name: DNA-directed RNA polymerase, mitochondrial / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO / EC number: DNA-directed RNA polymerase |
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Source (natural) | Organism: Homo sapiens (human) |
Molecular weight | Theoretical: 129.803094 KDa |
Recombinant expression | Organism: Escherichia coli BL21(DE3) (bacteria) |
Sequence | String: MGHHHHHHEN LYFQSNARKV QMGAKDATPV PCGRWAKILE KDKRTQQMRM QRLKAKLQMP FQSGEFKALT RRLQVEPRLL SKQMAGCLE DCTRQAPESP WEEQLARLLQ EAPGKLSLDV EQAPSGQHSQ AQLSGQQQRL LAFFKCCLLT DQLPLAHHLL V VHHGQRQK ...String: MGHHHHHHEN LYFQSNARKV QMGAKDATPV PCGRWAKILE KDKRTQQMRM QRLKAKLQMP FQSGEFKALT RRLQVEPRLL SKQMAGCLE DCTRQAPESP WEEQLARLLQ EAPGKLSLDV EQAPSGQHSQ AQLSGQQQRL LAFFKCCLLT DQLPLAHHLL V VHHGQRQK RKLLTLDMYN AVMLGWARQG AFKELVYVLF MVKDAGLTPD LLSYAAALQC MGRQDQDAGT IERCLEQMSQ EG LKLQALF TAVLLSEEDR ATVLKAVHKV KPTFSLPPQL PPPVNTSKLL RDVYAKDGRV SYPKLHLPLK TLQCLFEKQL HME LASRVC VVSVEKPTLP SKEVKHARKT LKTLRDQWEK ALCRALRETK NRLEREVYEG RFSLYPFLCL LDEREVVRML LQVL QALPA QGESFTTLAR ELSARTFSRH VVQRQRVSGQ VQALQNHYRK YLCLLASDAE VPEPCLPRQY WEALGAPEAL REQPW PLPV QMELGKLLAE MLVQATQMPC SLDKPHRSSR LVPVLYHVYS FRNVQQIGIL KPHPAYVQLL EKAAEPTLTF EAVDVP MLC PPLPWTSPHS GAFLLSPTKL MRTVEGATQH QELLETCPPT ALHGALDALT QLGNCAWRVN GRVLDLVLQL FQAKGCP QL GVPAPPSEAP QPPEAHLPHS AAPARKAELR RELAHCQKVA REMHSLRAEA LYRLSLAQHL RDRVFWLPHN MDFRGRTY P CPPHFNHLGS DVARALLEFA QGRPLGPHGL DWLKIHLVNL TGLKKREPLR KRLAFAEEVM DDILDSADQP LTGRKWWMG AEEPWQTLAC CMEVANAVRA SDPAAYVSHL PVHQDGSCNG LQHYAALGRD SVGAASVNLE PSDVPQDVYS GVAAQVEVFR RQDAQRGMR VAQVLEGFIT RKVVKQTVMT VVYGVTRYGG RLQIEKRLRE LSDFPQEFVW EASHYLVRQV FKSLQEMFSG T RAIQHWLT ESARLISHMG SVVEWVTPLG VPVIQPYRLD SKVKQIGGGI QSITYTHNGD ISRKPNTRKQ KNGFPPNFIH SL DSSHMML TALHCYRKGL TFVSVHDCYW THAADVSVMN QVCREQFVRL HSEPILQDLS RFLVKRFCSE PQKILEASQL KET LQAVPK PGAFDLEQVK RSTYFFS UniProtKB: DNA-directed RNA polymerase, mitochondrial |
-Macromolecule #2: (3~{R})-1-[(2~{R})-2-[4-(2-chloranyl-4-fluoranyl-phenyl)-2-oxidan...
Macromolecule | Name: (3~{R})-1-[(2~{R})-2-[4-(2-chloranyl-4-fluoranyl-phenyl)-2-oxidanylidene-chromen-7-yl]oxypropanoyl]piperidine-3-carboxylic acid type: ligand / ID: 2 / Number of copies: 1 / Formula: R4Q |
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Molecular weight | Theoretical: 473.878 Da |
Chemical component information | ChemComp-R4Q: |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Buffer | pH: 8 Component:
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Grid | Model: UltrAuFoil R2/2 / Material: GOLD / Pretreatment - Type: GLOW DISCHARGE | ||||||||||
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277.15 K / Instrument: FEI VITROBOT MARK IV |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Specialist optics | Energy filter - Name: GIF Bioquantum / Energy filter - Slit width: 20 eV |
Details | Stage was tilted 40 deg. |
Image recording | Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Average electron dose: 36.25 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Illumination mode: SPOT SCAN / Imaging mode: BRIGHT FIELD / Nominal defocus max: 3.0 µm / Nominal defocus min: 0.5 µm / Nominal magnification: 105000 |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |