+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-27689 | |||||||||
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Title | Sub-tomogram average of pro-meprin alpha supercoiled filament | |||||||||
Map data | Full reconstruction (unsharpened, masked, filtered to global 0.143 FSC) | |||||||||
Sample |
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Biological species | Homo sapiens (human) | |||||||||
Method | subtomogram averaging / cryo EM / Resolution: 12.7 Å | |||||||||
Authors | Bayly-Jones C / Lupton CJ / Fritz C / Schlenzig D / Whisstock JC | |||||||||
Funding support | Germany, Australia, 2 items
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Citation | Journal: Nat Commun / Year: 2022 Title: Helical ultrastructure of the metalloprotease meprin α in complex with a small molecule inhibitor. Authors: Charles Bayly-Jones / Christopher J Lupton / Claudia Fritz / Hariprasad Venugopal / Daniel Ramsbeck / Michael Wermann / Christian Jäger / Alex de Marco / Stephan Schilling / Dagmar ...Authors: Charles Bayly-Jones / Christopher J Lupton / Claudia Fritz / Hariprasad Venugopal / Daniel Ramsbeck / Michael Wermann / Christian Jäger / Alex de Marco / Stephan Schilling / Dagmar Schlenzig / James C Whisstock / Abstract: The zinc-dependent metalloprotease meprin α is predominantly expressed in the brush border membrane of proximal tubules in the kidney and enterocytes in the small intestine and colon. In normal ...The zinc-dependent metalloprotease meprin α is predominantly expressed in the brush border membrane of proximal tubules in the kidney and enterocytes in the small intestine and colon. In normal tissue homeostasis meprin α performs key roles in inflammation, immunity, and extracellular matrix remodelling. Dysregulated meprin α is associated with acute kidney injury, sepsis, urinary tract infection, metastatic colorectal carcinoma, and inflammatory bowel disease. Accordingly, meprin α is the target of drug discovery programs. In contrast to meprin β, meprin α is secreted into the extracellular space, whereupon it oligomerises to form giant assemblies and is the largest extracellular protease identified to date (~6 MDa). Here, using cryo-electron microscopy, we determine the high-resolution structure of the zymogen and mature form of meprin α, as well as the structure of the active form in complex with a prototype small molecule inhibitor and human fetuin-B. Our data reveal that meprin α forms a giant, flexible, left-handed helical assembly of roughly 22 nm in diameter. We find that oligomerisation improves proteolytic and thermal stability but does not impact substrate specificity or enzymatic activity. Furthermore, structural comparison with meprin β reveal unique features of the active site of meprin α, and helical assembly more broadly. | |||||||||
History |
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-Structure visualization
Supplemental images |
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-Downloads & links
-EMDB archive
Map data | emd_27689.map.gz | 2.4 MB | EMDB map data format | |
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Header (meta data) | emd-27689-v30.xml emd-27689.xml | 23.3 KB 23.3 KB | Display Display | EMDB header |
FSC (resolution estimation) | emd_27689_fsc.xml | 3.5 KB | Display | FSC data file |
Images | emd_27689.png | 76.9 KB | ||
Masks | emd_27689_msk_1.map | 3.4 MB | Mask map | |
Others | emd_27689_additional_1.map.gz emd_27689_additional_2.map.gz emd_27689_half_map_1.map.gz emd_27689_half_map_2.map.gz | 3.1 MB 741.9 KB 2.4 MB 2.4 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-27689 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-27689 | HTTPS FTP |
-Validation report
Summary document | emd_27689_validation.pdf.gz | 594.2 KB | Display | EMDB validaton report |
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Full document | emd_27689_full_validation.pdf.gz | 593.7 KB | Display | |
Data in XML | emd_27689_validation.xml.gz | 9.1 KB | Display | |
Data in CIF | emd_27689_validation.cif.gz | 11.8 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-27689 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-27689 | HTTPS FTP |
-Related structure data
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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-Map
File | Download / File: emd_27689.map.gz / Format: CCP4 / Size: 3.4 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||
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Annotation | Full reconstruction (unsharpened, masked, filtered to global 0.143 FSC) | ||||||||||||||||||||
Voxel size | X=Y=Z: 4.5 Å | ||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||
Details | EMDB XML:
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-Supplemental data
-Mask #1
File | emd_27689_msk_1.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Additional map: DeepEMhancer sharpened map (wide mode)
File | emd_27689_additional_1.map | ||||||||||||
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Annotation | DeepEMhancer sharpened map (wide mode) | ||||||||||||
Projections & Slices |
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Density Histograms |
-Additional map: B-factor sharpened map (Relion)
File | emd_27689_additional_2.map | ||||||||||||
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Annotation | B-factor sharpened map (Relion) | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: Half map 1 of 2
File | emd_27689_half_map_1.map | ||||||||||||
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Annotation | Half map 1 of 2 | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: Half map 2 of 2
File | emd_27689_half_map_2.map | ||||||||||||
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Annotation | Half map 2 of 2 | ||||||||||||
Projections & Slices |
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Density Histograms |
-Sample components
-Entire : Roughly 12 subunits of helical meprin alpha in the zymogen state
Entire | Name: Roughly 12 subunits of helical meprin alpha in the zymogen state |
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Components |
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-Supramolecule #1: Roughly 12 subunits of helical meprin alpha in the zymogen state
Supramolecule | Name: Roughly 12 subunits of helical meprin alpha in the zymogen state type: complex / Chimera: Yes / ID: 1 / Parent: 0 / Macromolecule list: all Details: Sub-tomogram average of a ~12 subunit region of recombinant, secreted helical pro-meprin alpha |
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Source (natural) | Organism: Homo sapiens (human) |
Recombinant expression | Organism: Drosophila (fruit flies) / Recombinant cell: Schneider-2 / Recombinant plasmid: pMT/BiP/V5 |
Molecular weight | Theoretical: 85 kDa/nm |
-Macromolecule #1: Pro-meprin alpha
Macromolecule | Name: Pro-meprin alpha / type: protein_or_peptide / ID: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: Homo sapiens (human) |
Recombinant expression | Organism: Drosophila melanogaster (fruit fly) |
Sequence | String: WSHPQFEKVP IKYLPEENVH DADFGEQKDI SEINLAAGLD LFQGDIL LQ KSRNGLRDPN TRWTFPIPYI LADNLGLNAK GAILYAFEMF RLKSCVDFKP YEGESSYI I FQQFDGCWSE VGDQHVGQNI SIGQGCAYKA IIEHEILHAL GFYHEQSRTD RDDYVNIWW ...String: WSHPQFEKVP IKYLPEENVH DADFGEQKDI SEINLAAGLD LFQGDIL LQ KSRNGLRDPN TRWTFPIPYI LADNLGLNAK GAILYAFEMF RLKSCVDFKP YEGESSYI I FQQFDGCWSE VGDQHVGQNI SIGQGCAYKA IIEHEILHAL GFYHEQSRTD RDDYVNIWW DQILSGYQHN FDTYDDSLIT DLNTPYDYES LMHYQPFSFN KNASVPTITA KIPEFNSIIG QRLDFSAID LERLNRMYNC TTTHTLLDHC TFEKANICGM IQGTRDDTDW AHQDSAQAGE V DHTLLGQC TGAGYFMQFS TSSGSAEEAA LLESRILYPK RKQQCLQFFY KMTGSPSDRL VV WVRRDDS TGNVRKLVKV QTFQGDDDHN WKIAHVVLKE EQKFRYLFQG TKGDPQNSTG GIY LDDITL TETPCPTGVW TVRNFSQVLE NTSKGDKLQS PRFYNSEGYG FGVTLYPNSR ESSG YLRLA FHVCSGENDA ILEWPVENRQ VIITILDQEP DVRNRMSSSM VFTTSKSHTS PAIND TVIW DRPSRVGTYH TDCNCFRSID LGWSGFISHQ MLKRRSFLKN DDLIIFVDFE DITHLS QTE VPTKGKRLSP QGLILQGQEQ QVSEEGSGKA MLEEALPVSL SQGQPSRQKR SVENTGP LE DHNWPQYFRD PCDPNPCQND GICVNVKGMA SCRCISGHAF FYTGERCQAV QVHGSVLG M VIGGTAGVIF LTFSIIAILS QRPRK |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | subtomogram averaging |
Aggregation state | filament |
-Sample preparation
Concentration | 0.5 mg/mL | |||||||||
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Buffer | pH: 7.4 Component:
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Grid | Model: Quantifoil R2/2 / Material: COPPER / Support film - Material: CARBON / Support film - topology: HOLEY | |||||||||
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277.15 K / Instrument: FEI VITROBOT MARK IV / Details: 3 s blot, -3 force. | |||||||||
Details | Polydisperse |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Image recording | Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Digitization - Frames/image: 1-40 / Number grids imaged: 1 / Average electron dose: 2.5 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 3.2 µm / Nominal defocus min: 2.5 µm / Nominal magnification: 64000 |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |