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- EMDB-12142: ASCT2 in the presence of the inhibitor Lc-BPE in the outward-open... -
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Open data
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Basic information
Entry | Database: EMDB / ID: EMD-12142 | ||||||||||||||||||||||||
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Title | ASCT2 in the presence of the inhibitor Lc-BPE in the outward-open conformation. | ||||||||||||||||||||||||
![]() | Cryo-EM map of ASCT2 in the presence of the inhibitor Lc-BPE at 3.43 A resolution in the outward-open conformation. Map was sharpened with a b factor of 189 A2 | ||||||||||||||||||||||||
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![]() | Solute carrier transporter / membrane protein / homology modeling / cancer metabolism / glutamine deprivation / cryo-EM | ||||||||||||||||||||||||
Function / homology | ![]() glutamine secretion / L-glutamine import across plasma membrane / glutamine transport / L-glutamine transmembrane transporter activity / L-serine transmembrane transporter activity / ligand-gated channel activity / neutral amino acid transport / amino acid transmembrane transporter activity / L-aspartate transmembrane transporter activity / Amino acid transport across the plasma membrane ...glutamine secretion / L-glutamine import across plasma membrane / glutamine transport / L-glutamine transmembrane transporter activity / L-serine transmembrane transporter activity / ligand-gated channel activity / neutral amino acid transport / amino acid transmembrane transporter activity / L-aspartate transmembrane transporter activity / Amino acid transport across the plasma membrane / L-aspartate import across plasma membrane / neutral L-amino acid transmembrane transporter activity / symporter activity / antiporter activity / amino acid transport / RHOJ GTPase cycle / RHOQ GTPase cycle / protein homotrimerization / RHOH GTPase cycle / transport across blood-brain barrier / RAC3 GTPase cycle / RAC1 GTPase cycle / erythrocyte differentiation / basal plasma membrane / melanosome / virus receptor activity / signaling receptor activity / extracellular exosome / membrane / metal ion binding / plasma membrane Similarity search - Function | ||||||||||||||||||||||||
Biological species | ![]() | ||||||||||||||||||||||||
Method | single particle reconstruction / cryo EM / Resolution: 3.43 Å | ||||||||||||||||||||||||
![]() | Garibsingh RA / Ndaru E | ||||||||||||||||||||||||
Funding support | ![]() ![]()
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![]() | ![]() Title: Rational design of ASCT2 inhibitors using an integrated experimental-computational approach. Authors: Rachel-Ann A Garibsingh / Elias Ndaru / Alisa A Garaeva / Yueyue Shi / Laura Zielewicz / Paul Zakrepine / Massimiliano Bonomi / Dirk J Slotboom / Cristina Paulino / Christof Grewer / Avner Schlessinger / ![]() ![]() ![]() Abstract: ASCT2 (SLC1A5) is a sodium-dependent neutral amino acid transporter that controls amino acid homeostasis in peripheral tissues. In cancer, ASCT2 is up-regulated where it modulates intracellular ...ASCT2 (SLC1A5) is a sodium-dependent neutral amino acid transporter that controls amino acid homeostasis in peripheral tissues. In cancer, ASCT2 is up-regulated where it modulates intracellular glutamine levels, fueling cell proliferation. Nutrient deprivation via ASCT2 inhibition provides a potential strategy for cancer therapy. Here, we rationally designed stereospecific inhibitors exploiting specific subpockets in the substrate binding site using computational modeling and cryo-electron microscopy (cryo-EM). The final structures combined with molecular dynamics simulations reveal multiple pharmacologically relevant conformations in the ASCT2 binding site as well as a previously unknown mechanism of stereospecific inhibition. Furthermore, this integrated analysis guided the design of a series of unique ASCT2 inhibitors. Our results provide a framework for future development of cancer therapeutics targeting nutrient transport via ASCT2, as well as demonstrate the utility of combining computational modeling and cryo-EM for solute carrier ligand discovery. | ||||||||||||||||||||||||
History |
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Structure visualization
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Structure viewer | EM map: ![]() ![]() ![]() |
Supplemental images |
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Downloads & links
-EMDB archive
Map data | ![]() | 28.5 MB | ![]() | |
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Header (meta data) | ![]() ![]() | 20.7 KB 20.7 KB | Display Display | ![]() |
Images | ![]() | 70.6 KB | ||
Masks | ![]() | 30.5 MB | ![]() | |
Filedesc metadata | ![]() | 6.7 KB | ||
Others | ![]() ![]() | 23.1 MB 23.1 MB | ||
Archive directory | ![]() ![]() | HTTPS FTP |
-Validation report
Summary document | ![]() | 745.3 KB | Display | ![]() |
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Full document | ![]() | 744.9 KB | Display | |
Data in XML | ![]() | 10.6 KB | Display | |
Data in CIF | ![]() | 12.3 KB | Display | |
Arichive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 7bcqMC ![]() 7bcsMC ![]() 7bctC M: atomic model generated by this map C: citing same article ( |
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Similar structure data |
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Links
EMDB pages | ![]() ![]() |
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Related items in Molecule of the Month |
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Map
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Annotation | Cryo-EM map of ASCT2 in the presence of the inhibitor Lc-BPE at 3.43 A resolution in the outward-open conformation. Map was sharpened with a b factor of 189 A2 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 1.012 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
-Mask #1
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Density Histograms |
-Half map: half-map 1 used for post processing step and...
File | emd_12142_half_map_1.map | ||||||||||||
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Annotation | half-map 1 used for post processing step and FSC resolution calculation. | ||||||||||||
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Density Histograms |
-Half map: half-map 2 used for post processing step and...
File | emd_12142_half_map_2.map | ||||||||||||
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Annotation | half-map 2 used for post processing step and FSC resolution calculation. | ||||||||||||
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Density Histograms |
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Sample components
-Entire : ASCT2 in the presence of the inhibitor Lc-BPE in the outward-open...
Entire | Name: ASCT2 in the presence of the inhibitor Lc-BPE in the outward-open conformation. |
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Components |
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-Supramolecule #1: ASCT2 in the presence of the inhibitor Lc-BPE in the outward-open...
Supramolecule | Name: ASCT2 in the presence of the inhibitor Lc-BPE in the outward-open conformation. type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1 |
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Source (natural) | Organism: ![]() |
-Macromolecule #1: Neutral amino acid transporter B(0)
Macromolecule | Name: Neutral amino acid transporter B(0) / type: protein_or_peptide / ID: 1 / Number of copies: 3 / Enantiomer: LEVO |
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Source (natural) | Organism: ![]() |
Molecular weight | Theoretical: 56.638902 KDa |
Recombinant expression | Organism: ![]() |
Sequence | String: MVADPPRDSK GLAAAEPTAN GGLALASIED QGAAAGGYCG SRDQVRRCLR ANLLVLLTVV AVVAGVALGL GVSGAGGALA LGPERLSAF VFPGELLLRL LRMIILPLVV CSLIGGAASL DPGALGRLGA WALLFFLVTT LLASALGVGL ALALQPGAAS A AINASVGA ...String: MVADPPRDSK GLAAAEPTAN GGLALASIED QGAAAGGYCG SRDQVRRCLR ANLLVLLTVV AVVAGVALGL GVSGAGGALA LGPERLSAF VFPGELLLRL LRMIILPLVV CSLIGGAASL DPGALGRLGA WALLFFLVTT LLASALGVGL ALALQPGAAS A AINASVGA AGSAENAPSK EVLDSFLDLA RNIFPSNLVS AAFRSYSTTY EERNITGTRV KVPVGQEVEG MNILGLVVFA IV FGVALRK LGPEGELLIR FFNSFNEATM VLVSWIMWYA PVGIMFLVAG KIVEMEDVGL LFARLGKYIL CCLLGHAIHG LLV LPLIYF LFTRKNPYRF LWGIVTPLAT AFGTSSSSAT LPLMMKCVEE NNGVAKHISR FILPIGATVN MDGAALFQCV AAVF IAQLS QQSLDFVKII TILVTATASS VGAAGIPAGG VLTLAIILEA VNLPVDHISL ILAVDWLVDR SCTVLNVEGD ALGAG LLQN YVDRTESRST EPELIQVKSE LPLDPLPVPT EEGNPLLKHY RGPAGDATVA SEKESVM UniProtKB: Neutral amino acid transporter B(0) |
-Macromolecule #2: 4-(4-phenylphenyl)carbonyloxypyrrolidine-2-carboxylic acid
Macromolecule | Name: 4-(4-phenylphenyl)carbonyloxypyrrolidine-2-carboxylic acid type: ligand / ID: 2 / Number of copies: 3 / Formula: TG2 |
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Molecular weight | Theoretical: 311.332 Da |
Chemical component information | ![]() ChemComp-TG2: |
-Experimental details
-Structure determination
Method | cryo EM |
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![]() | single particle reconstruction |
Aggregation state | particle |
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Sample preparation
Concentration | 1 mg/mL |
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Buffer | pH: 7.4 / Details: 20 mM Tris-HCl, pH 7.4, 200 mM NaCl |
Grid | Model: Quantifoil R1.2/1.3 / Material: GOLD / Mesh: 300 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 30 sec. / Details: at 5mA |
Vitrification | Cryogen name: ETHANE-PROPANE / Chamber humidity: 100 % / Chamber temperature: 288 K / Instrument: FEI VITROBOT MARK IV |
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Electron microscopy
Microscope | FEI TALOS ARCTICA |
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Temperature | Min: 90.0 K / Max: 105.0 K |
Specialist optics | Energy filter - Name: GIF Bioquantum / Energy filter - Slit width: 20 eV |
Image recording | Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Digitization - Dimensions - Width: 3838 pixel / Digitization - Dimensions - Height: 3710 pixel / Digitization - Frames/image: 1-60 / Number grids imaged: 3 / Number real images: 6233 / Average exposure time: 9.0 sec. / Average electron dose: 53.0 e/Å2 |
Electron beam | Acceleration voltage: 200 kV / Electron source: ![]() |
Electron optics | C2 aperture diameter: 100.0 µm / Calibrated defocus max: 2.0 µm / Calibrated defocus min: 0.3 µm / Calibrated magnification: 49407 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 2.0 µm / Nominal defocus min: 0.3 µm / Nominal magnification: 130000 |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN |
Experimental equipment | ![]() Model: Talos Arctica / Image courtesy: FEI Company |
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Image processing
-Atomic model buiding 1
Initial model | PDB ID: Chain - Source name: PDB / Chain - Initial model type: experimental model |
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Refinement | Space: REAL |
Output model | ![]() PDB-7bcq: ![]() PDB-7bcs: |