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Yorodumi- EMDB-14435: Abortive infection DNA polymerase AbiK from Lactococcus lactis, Y... -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-14435 | |||||||||
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Title | Abortive infection DNA polymerase AbiK from Lactococcus lactis, Y44F variant | |||||||||
Map data | ||||||||||
Sample |
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Keywords | DNA polymerase / protein-primed DNA synthesis / template-independent DNA synthesis / abortive infection / ANTIVIRAL PROTEIN | |||||||||
Function / homology | Reverse transcriptase domain / Reverse transcriptase (RT) catalytic domain profile. / Reverse transcriptase (RNA-dependent DNA polymerase) / DNA/RNA polymerase superfamily / metal ion binding / AbiK Function and homology information | |||||||||
Biological species | Lactococcus lactis (lactic acid bacteria) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 2.68 Å | |||||||||
Authors | Figiel M / Gapinska M / Czarnocki-Cieciura M / Zajko W / Nowotny M | |||||||||
Funding support | Poland, European Union, 2 items
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Citation | Journal: Nucleic Acids Res / Year: 2022 Title: Mechanism of protein-primed template-independent DNA synthesis by Abi polymerases. Authors: Małgorzata Figiel / Marta Gapińska / Mariusz Czarnocki-Cieciura / Weronika Zajko / Małgorzata Sroka / Krzysztof Skowronek / Marcin Nowotny Abstract: Abortive infection (Abi) is a bacterial antiphage defense strategy involving suicide of the infected cell. Some Abi pathways involve polymerases that are related to reverse transcriptases. They are ...Abortive infection (Abi) is a bacterial antiphage defense strategy involving suicide of the infected cell. Some Abi pathways involve polymerases that are related to reverse transcriptases. They are unique in the way they combine the ability to synthesize DNA in a template-independent manner with protein priming. Here, we report crystal and cryo-electron microscopy structures of two Abi polymerases: AbiK and Abi-P2. Both proteins adopt a bilobal structure with an RT-like domain that comprises palm and fingers subdomains and a unique helical domain. AbiK and Abi-P2 adopt a hexameric and trimeric configuration, respectively, which is unprecedented for reverse transcriptases. Biochemical experiments showed that the formation of these oligomers is required for the DNA polymerization activity. The structure of the AbiK-DNA covalent adduct visualized interactions between the 3' end of DNA and the active site and covalent attachment of the 5' end of DNA to a tyrosine residue used for protein priming. Our data reveal a structural basis of the mechanism of highly unusual template-independent protein-priming polymerases. | |||||||||
History |
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-Structure visualization
Supplemental images |
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-Downloads & links
-EMDB archive
Map data | emd_14435.map.gz | 8.9 MB | EMDB map data format | |
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Header (meta data) | emd-14435-v30.xml emd-14435.xml | 17 KB 17 KB | Display Display | EMDB header |
FSC (resolution estimation) | emd_14435_fsc.xml | 12.6 KB | Display | FSC data file |
Images | emd_14435.png | 164.8 KB | ||
Masks | emd_14435_msk_1.map | 216 MB | Mask map | |
Filedesc metadata | emd-14435.cif.gz | 6.1 KB | ||
Others | emd_14435_half_map_1.map.gz emd_14435_half_map_2.map.gz | 200.1 MB 200.1 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-14435 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-14435 | HTTPS FTP |
-Validation report
Summary document | emd_14435_validation.pdf.gz | 887.3 KB | Display | EMDB validaton report |
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Full document | emd_14435_full_validation.pdf.gz | 886.9 KB | Display | |
Data in XML | emd_14435_validation.xml.gz | 21.7 KB | Display | |
Data in CIF | emd_14435_validation.cif.gz | 28.2 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-14435 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-14435 | HTTPS FTP |
-Related structure data
Related structure data | 7z0zMC 7r06C 7r07C 7r08C 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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-Map
File | Download / File: emd_14435.map.gz / Format: CCP4 / Size: 216 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||
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Projections & slices | Image control
Images are generated by Spider. | ||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 0.82 Å | ||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
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-Supplemental data
-Mask #1
File | emd_14435_msk_1.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Half map: #2
File | emd_14435_half_map_1.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Half map: #1
File | emd_14435_half_map_2.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Sample components
-Entire : Homohexamer of L. lactis AbiK Y44F mutant
Entire | Name: Homohexamer of L. lactis AbiK Y44F mutant |
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Components |
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-Supramolecule #1: Homohexamer of L. lactis AbiK Y44F mutant
Supramolecule | Name: Homohexamer of L. lactis AbiK Y44F mutant / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all |
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Source (natural) | Organism: Lactococcus lactis (lactic acid bacteria) |
Molecular weight | Theoretical: 420 KDa |
-Macromolecule #1: AbiK
Macromolecule | Name: AbiK / type: protein_or_peptide / ID: 1 / Number of copies: 6 / Enantiomer: LEVO |
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Source (natural) | Organism: Lactococcus lactis (lactic acid bacteria) |
Molecular weight | Theoretical: 71.617109 KDa |
Recombinant expression | Organism: Escherichia coli BL21 (bacteria) |
Sequence | String: GSMKKEFTEL YDFIFDPIFL VRYGYYDRSI KNKKMNTAKV ELDNEFGKSD SFYFKVFNME SFADYLRSHD LKTHFNGKKP LSTDPVYFN IPKNIEARRQ YKMPNLYSYM ALNYYICDNK KEFIEVFIDN KFSTSKFFNQ LNFDYPKTQE ITQTLLYGGI K KLHLDLSN ...String: GSMKKEFTEL YDFIFDPIFL VRYGYYDRSI KNKKMNTAKV ELDNEFGKSD SFYFKVFNME SFADYLRSHD LKTHFNGKKP LSTDPVYFN IPKNIEARRQ YKMPNLYSYM ALNYYICDNK KEFIEVFIDN KFSTSKFFNQ LNFDYPKTQE ITQTLLYGGI K KLHLDLSN FYHTLYTHSI PWMIDGKSAS KQNRKKGFSN TLDTLITACQ YDETHGIPTG NLLSRIITEL YMCHFDKQME YK KFVYSRY VDDFIFPFTF ENEKQEFLNE FNLICRENNL IINDNKTKVD NFPFVDKSSK SDIFSFFENI TSTNSNDKWI KEI SNFIDY CVNEEHLGNK GAIKCIFPVI TNTLKQKKVD TKNIDNIFSK RNMVTNFNVF EKILDLSLKD SRLTNKFLTF FENI NEFGF SSLSASNIVK KYFSNNSKGL KEKIDHYRKN NFNQELYQIL LYMVVFEIDD LLNQEELLNL IDLNIDDYSL ILGTI LYLK NSSYKLEKLL KKIDQLFINT HANYDVKTSR MAEKLWLFRY FFYFLNCKNI FSQKEINSYC QSQNYNSGQN GYQTEL NWN YIKGQGKDLR ANNFFNELIV KEVWLISCGE NEDFKYLN UniProtKB: AbiK |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 1.74 mg/mL | |||||||||||||||
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Buffer | pH: 7 Component:
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Grid | Model: C-flat-2/1 / Material: COPPER / Mesh: 200 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 60 sec. | |||||||||||||||
Vitrification | Cryogen name: ETHANE / 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 |
Image recording | Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Average electron dose: 60.0 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.5 µm / Nominal defocus min: 1.5 µm |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |