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Yorodumi- EMDB-10735: Cryo-EM structure of trimeric human STEAP1 bound to three Fab120.... -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-10735 | |||||||||
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Title | Cryo-EM structure of trimeric human STEAP1 bound to three Fab120.545 fragments | |||||||||
Map data | Unsharpened cryo-EM map of homotrimeric STEAP1 bound to three Fab-fragments of antibody mAb120.545. The constant region of the Fabs is masked out. | |||||||||
Sample |
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Function / homology | Function and homology information cell-cell junction / endosome membrane / endosome / heme binding / membrane / metal ion binding / plasma membrane Similarity search - Function | |||||||||
Biological species | Homo sapiens (human) / house mouse (house mouse) / Mus musculus (house mouse) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 2.97 Å | |||||||||
Authors | Oosterheert W / Gros P | |||||||||
Funding support | Netherlands, 1 items
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Citation | Journal: J Biol Chem / Year: 2020 Title: Cryo-electron microscopy structure and potential enzymatic function of human six-transmembrane epithelial antigen of the prostate 1 (STEAP1). Authors: Wout Oosterheert / Piet Gros / Abstract: Six-transmembrane epithelial antigen of the prostate 1 (STEAP1) is an integral membrane protein that is highly up-regulated on the cell surface of several human cancers, making it a promising ...Six-transmembrane epithelial antigen of the prostate 1 (STEAP1) is an integral membrane protein that is highly up-regulated on the cell surface of several human cancers, making it a promising therapeutic target to manage these diseases. It shares sequence homology with three enzymes (STEAP2-STEAP4) that catalyze the NADPH-dependent reduction of iron(III). However, STEAP1 lacks an intracellular NADPH-binding domain and does not exhibit cellular ferric reductase activity. Thus, both the molecular function of STEAP1 and its role in cancer progression remain elusive. Here, we present a ∼3.0-Å cryo-EM structure of trimeric human STEAP1 bound to three antigen-binding fragments (Fabs) of the clinically used antibody mAb120.545. The structure revealed that STEAP1 adopts a reductase-like conformation and interacts with the Fabs through its extracellular helices. Enzymatic assays in human cells revealed that STEAP1 promotes iron(III) reduction when fused to the intracellular NADPH-binding domain of its family member STEAP4, suggesting that STEAP1 functions as a ferric reductase in STEAP heterotrimers. Our work provides a foundation for deciphering the molecular mechanisms of STEAP1 and may be useful in the design of new therapeutic strategies to target STEAP1 in cancer. | |||||||||
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_10735.map.gz | 4.4 MB | EMDB map data format | |
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Header (meta data) | emd-10735-v30.xml emd-10735.xml | 32.2 KB 32.2 KB | Display Display | EMDB header |
FSC (resolution estimation) | emd_10735_fsc.xml | 10.7 KB | Display | FSC data file |
Images | emd_10735.png | 107.2 KB | ||
Masks | emd_10735_msk_1.map | 103 MB | Mask map | |
Others | emd_10735_additional_1.map.gz emd_10735_additional_2.map.gz emd_10735_half_map_1.map.gz emd_10735_half_map_2.map.gz | 79.9 MB 5.7 MB 80.6 MB 80.6 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-10735 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-10735 | HTTPS FTP |
-Validation report
Summary document | emd_10735_validation.pdf.gz | 382.3 KB | Display | EMDB validaton report |
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Full document | emd_10735_full_validation.pdf.gz | 381.4 KB | Display | |
Data in XML | emd_10735_validation.xml.gz | 16.4 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-10735 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-10735 | HTTPS FTP |
-Related structure data
Related structure data | 6y9bMC 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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-Map
File | Download / File: emd_10735.map.gz / Format: CCP4 / Size: 103 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | Unsharpened cryo-EM map of homotrimeric STEAP1 bound to three Fab-fragments of antibody mAb120.545. The constant region of the Fabs is masked out. | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 1.0285 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_10735_msk_1.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Additional map: Unsharpened, unmasked cryo-EM map of homotrimeric STEAP1 bound...
File | emd_10735_additional_1.map | ||||||||||||
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Annotation | Unsharpened, unmasked cryo-EM map of homotrimeric STEAP1 bound to three Fab-fragments of antibody mAb120.545. | ||||||||||||
Projections & Slices |
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Density Histograms |
-Additional map: Sharpened cryo-EM map of homotrimeric STEAP1 bound to...
File | emd_10735_additional_2.map | ||||||||||||
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Annotation | Sharpened cryo-EM map of homotrimeric STEAP1 bound to three Fab-fragments of antibody mAb120.545. The constant region of the Fabs is masked out. The map is autosharpened (B = -69 Angstrom) in Relion. | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: Half map 1 of homotrimeric STEAP1 bound to...
File | emd_10735_half_map_1.map | ||||||||||||
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Annotation | Half map 1 of homotrimeric STEAP1 bound to three Fab-fragments of antibody mAb120.545. | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: Half map 2 of homotrimeric STEAP1 bound to...
File | emd_10735_half_map_2.map | ||||||||||||
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Annotation | Half map 2 of homotrimeric STEAP1 bound to three Fab-fragments of antibody mAb120.545. | ||||||||||||
Projections & Slices |
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Density Histograms |
-Sample components
+Entire : Trimeric human six-transmembrane epithelial antigen of the prosta...
+Supramolecule #1: Trimeric human six-transmembrane epithelial antigen of the prosta...
+Supramolecule #2: Fab-fragment 1 of antibody mAb120.545
+Supramolecule #3: Homo-trimeric STEAP1, chain a, b and c.
+Supramolecule #4: Fab Fragment 2 of antibody mAb120.545
+Supramolecule #5: Fab-fragment 3 of antibody mAb120.545
+Macromolecule #1: Metalloreductase STEAP1
+Macromolecule #2: Fab120.545 light chain
+Macromolecule #3: Fab120.545 heavy chain
+Macromolecule #4: PROTOPORPHYRIN IX CONTAINING FE
+Macromolecule #5: 1,2-DIMYRISTOYL-SN-GLYCERO-3-PHOSPHATE
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 5.0 mg/mL | |||||||||||||||
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Buffer | pH: 7.8 Component:
Details: Protein sample was subjected to size-exclusion chromatography in 20 mM Tris pH 7.8, 200 mM NaCl, 0.08% (w/v) digitonin. 1 mM FAD (final concentration) was added to the sample before vitrification. | |||||||||||||||
Grid | Model: Quantifoil R1.2/1.3 / Material: GOLD / Mesh: 200 / Pretreatment - Type: GLOW DISCHARGE | |||||||||||||||
Vitrification | Cryogen name: ETHANE-PROPANE / Chamber humidity: 100 % / Chamber temperature: 293 K / Instrument: FEI VITROBOT MARK IV / Details: blot 4 seconds, blotforce 0. | |||||||||||||||
Details | STEAP1-Fab120.545 complex purified in digitonin. The sample was monodisperse. |
-Electron microscopy
Microscope | FEI TALOS ARCTICA |
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Specialist optics | Energy filter - Name: GIF Quantum LS / Energy filter - Slit width: 20 eV |
Details | Data was collected on a 200 kV Talos Arctica, located at Utrecht University, the Netherlands. |
Image recording | Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: SUPER-RESOLUTION / Digitization - Frames/image: 1-26 / Number grids imaged: 1 / Number real images: 5325 / Average exposure time: 6.5 sec. / Average electron dose: 49.5 e/Å2 |
Electron beam | Acceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN |
Electron optics | C2 aperture diameter: 50.0 µm / Calibrated defocus max: 3.5 µm / Calibrated defocus min: 0.4 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 3.0 µm / Nominal defocus min: 1.0 µm / Nominal magnification: 130000 |
Sample stage | Cooling holder cryogen: NITROGEN |
Experimental equipment | Model: Talos Arctica / Image courtesy: FEI Company |