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Yorodumi- EMDB-41959: Original Cryo-EM map of the SPARTA oligomer with guide RNA and ta... -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-41959 | |||||||||
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Title | Original Cryo-EM map of the SPARTA oligomer with guide RNA and target DNA | |||||||||
Map data | raw consensus map | |||||||||
Sample |
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Keywords | RNA / DNA / Argonaute / TIR / immune system / RNA BINDING PROTEIN | |||||||||
Biological species | Thermoflavifilum thermophilum (bacteria) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 3.35 Å | |||||||||
Authors | Malik R / Kottur J / Aggarwal AK | |||||||||
Funding support | United States, 1 items
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Citation | Journal: Nat Commun / Year: 2024 Title: Nucleic acid mediated activation of a short prokaryotic Argonaute immune system. Authors: Jithesh Kottur / Radhika Malik / Aneel K Aggarwal / Abstract: A short prokaryotic Argonaute (pAgo) TIR-APAZ (SPARTA) defense system, activated by invading DNA to unleash its TIR domain for NAD(P) hydrolysis, was recently identified in bacteria. We report the ...A short prokaryotic Argonaute (pAgo) TIR-APAZ (SPARTA) defense system, activated by invading DNA to unleash its TIR domain for NAD(P) hydrolysis, was recently identified in bacteria. We report the crystal structure of SPARTA heterodimer in the absence of guide-RNA/target-ssDNA (2.66 Å) and a cryo-EM structure of the SPARTA oligomer (tetramer of heterodimers) bound to guide-RNA/target-ssDNA at nominal 3.15-3.35 Å resolution. The crystal structure provides a high-resolution view of SPARTA, revealing the APAZ domain as equivalent to the N, L1, and L2 regions of long pAgos and the MID domain containing a unique insertion (insert57). Cryo-EM structure reveals regions of the PIWI (loop10-9) and APAZ (helix αN) domains that reconfigure for nucleic-acid binding and decrypts regions/residues that reorganize to expose a positively charged pocket for higher-order assembly. The TIR domains amass in a parallel-strands arrangement for catalysis. We visualize SPARTA before and after RNA/ssDNA binding and uncover the basis of its active assembly leading to abortive infection. | |||||||||
History |
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-Structure visualization
Supplemental images |
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-Downloads & links
-EMDB archive
Map data | emd_41959.map.gz | 398.4 MB | EMDB map data format | |
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Header (meta data) | emd-41959-v30.xml emd-41959.xml | 14.8 KB 14.8 KB | Display Display | EMDB header |
Images | emd_41959.png | 138.9 KB | ||
Filedesc metadata | emd-41959.cif.gz | 5.1 KB | ||
Others | emd_41959_half_map_1.map.gz emd_41959_half_map_2.map.gz | 390.8 MB 390.8 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-41959 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-41959 | HTTPS FTP |
-Validation report
Summary document | emd_41959_validation.pdf.gz | 889.8 KB | Display | EMDB validaton report |
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Full document | emd_41959_full_validation.pdf.gz | 889.4 KB | Display | |
Data in XML | emd_41959_validation.xml.gz | 18.1 KB | Display | |
Data in CIF | emd_41959_validation.cif.gz | 21.4 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-41959 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-41959 | HTTPS FTP |
-Related structure data
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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-Map
File | Download / File: emd_41959.map.gz / Format: CCP4 / Size: 421.9 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||
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Annotation | raw consensus map | ||||||||||||||||||||
Voxel size | X=Y=Z: 1.083 Å | ||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||
Details | EMDB XML:
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-Supplemental data
-Half map: half-map
File | emd_41959_half_map_1.map | ||||||||||||
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Annotation | half-map | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: half-map
File | emd_41959_half_map_2.map | ||||||||||||
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Annotation | half-map | ||||||||||||
Projections & Slices |
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Density Histograms |
-Sample components
-Entire : SPARTA complex with gRNA/tDNA
Entire | Name: SPARTA complex with gRNA/tDNA |
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Components |
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-Supramolecule #1: SPARTA complex with gRNA/tDNA
Supramolecule | Name: SPARTA complex with gRNA/tDNA / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all |
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Source (natural) | Organism: Thermoflavifilum thermophilum (bacteria) |
-Macromolecule #1: pAgo
Macromolecule | Name: pAgo / type: protein_or_peptide / ID: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: Thermoflavifilum thermophilum (bacteria) |
Recombinant expression | Organism: Escherichia coli (E. coli) |
Sequence | String: MKELIYIEEP SILFAHGQKC TDPRDGLALF GPLNQIYGIK SGVVGTQKGL QIFKSYLDKI QKPIYNHNN ITRPMFPGFE AVFGCKWESQ NIVFKEITDE EIRRYLFNAS THKRTYDLVT L FNDKIITA NKNDEERVDV WFVIVPEEIY KYCRPNSVLP NELVQTKSLI ...String: MKELIYIEEP SILFAHGQKC TDPRDGLALF GPLNQIYGIK SGVVGTQKGL QIFKSYLDKI QKPIYNHNN ITRPMFPGFE AVFGCKWESQ NIVFKEITDE EIRRYLFNAS THKRTYDLVT L FNDKIITA NKNDEERVDV WFVIVPEEIY KYCRPNSVLP NELVQTKSLI SKSKAKSFRY TP TLFEEFN KKLKEVEKEA KTYNYDAQFH DQLKARLLEH TIPTQILRES TLAWRDFKNT FGA PIRDFS KIEGHLAWTI STAAYYKAGG KPWKLGDIRP GVCYLGLVYK KIEKSKNPQN ACCA AQMFL DNGDGTVFKG EVGPWYNPEK GEYHLKPKEA KALLTQALES YKEQNKSYPK EVFIH ARTR FNDEEWNAFN EVTPKNTNLV GVTITKSKPL KLYKTEGAFP IMRGNAYIVD EKKAFL WTL GFVPKLQSTL SMEVPNPIFI EINKGEAEIQ QVLKDILALT KLNYNACIYA DGEPVTL RF ANKIGEILTA STEIKTPPLA FKYYI |
-Macromolecule #2: TIR-APAZ
Macromolecule | Name: TIR-APAZ / type: protein_or_peptide / ID: 2 / Enantiomer: LEVO |
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Source (natural) | Organism: Thermoflavifilum thermophilum (bacteria) |
Recombinant expression | Organism: Escherichia coli (E. coli) |
Sequence | String: MRNKIFISHA TPEDDDFTRW LSLKLIGLGY EVWCDILFLD KGVDFWSTIE KEIRENTCKF LIVSSTAGN KREGVLKELA VATKVKKHLQ DDMFIIPLAI DENLSYDDIN IEIVRLNAID F KKSWAKGL QDLLDAFEKQ NVPKKPPDHS KSNLLYQQIF LHDKQAIEKE ...String: MRNKIFISHA TPEDDDFTRW LSLKLIGLGY EVWCDILFLD KGVDFWSTIE KEIRENTCKF LIVSSTAGN KREGVLKELA VATKVKKHLQ DDMFIIPLAI DENLSYDDIN IEIVRLNAID F KKSWAKGL QDLLDAFEKQ NVPKKPPDHS KSNLLYQQIF LHDKQAIEKE ETYDSNWFPI IS FPNELRF HRYDWRLPKQ FDVRTLAFPA IRYKEYLCTF AWEYDFIHQL PKTETYNGQE SIR ISTSDI LSGRYDTDFI RNYECQRLIV QLINKAFELR MKDKNVREYQ MSKTFAYWIE KGKL EKDKF EKIKLVGKQK NKYWHFGISA AGKLYPSPVL MVSSHIIFTM DGINLIKSKS IQHSS RRKQ GKNWWNDKWR EKLLAFIRFL SDDQNAIYLN VGSEEKILIS NKPLKFFGKM SYVTPS EVT LEEESVLADI NNFEEDTEDL DELEDIE |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Buffer | pH: 7.5 |
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Sugar embedding | Material: Vitreous ice |
Vitrification | Cryogen name: ETHANE |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Image recording | Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Average electron dose: 51.11 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: 0.8 µm |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
-Image processing
Startup model | Type of model: NONE |
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Final reconstruction | Resolution.type: BY AUTHOR / Resolution: 3.35 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 238432 |
Initial angle assignment | Type: MAXIMUM LIKELIHOOD |
Final angle assignment | Type: MAXIMUM LIKELIHOOD |