+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-28596 | |||||||||
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Title | CryoEM Structure of NLRP3 NACHT domain in complex with G2394 | |||||||||
Map data | ||||||||||
Sample |
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Keywords | NLRP3 / NACHT / inhibitor / HYDROLASE | |||||||||
Function / homology | Function and homology information small molecule sensor activity / detection of biotic stimulus / cysteine-type endopeptidase activator activity / phosphatidylinositol phosphate binding / positive regulation of T-helper 2 cell differentiation / NLRP3 inflammasome complex assembly / interphase microtubule organizing center / positive regulation of T-helper 2 cell cytokine production / NLRP3 inflammasome complex / positive regulation of type 2 immune response ...small molecule sensor activity / detection of biotic stimulus / cysteine-type endopeptidase activator activity / phosphatidylinositol phosphate binding / positive regulation of T-helper 2 cell differentiation / NLRP3 inflammasome complex assembly / interphase microtubule organizing center / positive regulation of T-helper 2 cell cytokine production / NLRP3 inflammasome complex / positive regulation of type 2 immune response / osmosensory signaling pathway / peptidoglycan binding / negative regulation of non-canonical NF-kappaB signal transduction / phosphatidylinositol-4-phosphate binding / pattern recognition receptor signaling pathway / microtubule organizing center / negative regulation of interleukin-1 beta production / positive regulation of interleukin-4 production / pyroptotic inflammatory response / negative regulation of acute inflammatory response / The NLRP3 inflammasome / protein maturation / Purinergic signaling in leishmaniasis infection / signaling adaptor activity / molecular condensate scaffold activity / positive regulation of interleukin-1 beta production / Hydrolases; Acting on acid anhydrides; Acting on acid anhydrides to facilitate cellular and subcellular movement / protein homooligomerization / Cytoprotection by HMOX1 / cellular response to virus / Metalloprotease DUBs / ADP binding / defense response / negative regulation of inflammatory response / positive regulation of inflammatory response / positive regulation of non-canonical NF-kappaB signal transduction / SARS-CoV-1 activates/modulates innate immune responses / positive regulation of NF-kappaB transcription factor activity / protein-macromolecule adaptor activity / cellular response to lipopolysaccharide / DNA-binding transcription factor binding / sequence-specific DNA binding / molecular adaptor activity / inflammatory response / Golgi membrane / innate immune response / apoptotic process / SARS-CoV-2 activates/modulates innate and adaptive immune responses / signal transduction / endoplasmic reticulum / positive regulation of transcription by RNA polymerase II / ATP hydrolysis activity / mitochondrion / extracellular region / ATP binding / identical protein binding / membrane / nucleus / cytosol / cytoplasm Similarity search - Function | |||||||||
Biological species | Homo sapiens (human) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 3.5 Å | |||||||||
Authors | Murray JM / Johnson MC | |||||||||
Funding support | 1 items
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Citation | Journal: J Med Chem / Year: 2022 Title: Overcoming Preclinical Safety Obstacles to Discover ()--((1,2,3,5,6,7-Hexahydro--indacen-4-yl)carbamoyl)-6-(methylamino)-6,7-dihydro-5-pyrazolo[5,1-][1,3]oxazine-3-sulfonamide (GDC-2394): A ...Title: Overcoming Preclinical Safety Obstacles to Discover ()--((1,2,3,5,6,7-Hexahydro--indacen-4-yl)carbamoyl)-6-(methylamino)-6,7-dihydro-5-pyrazolo[5,1-][1,3]oxazine-3-sulfonamide (GDC-2394): A Potent and Selective NLRP3 Inhibitor. Authors: Christopher McBride / Lynnie Trzoss / Davide Povero / Milos Lazic / Geza Ambrus-Aikelin / Angelina Santini / Rama Pranadinata / Gretchen Bain / Ryan Stansfield / Jeffrey A Stafford / James ...Authors: Christopher McBride / Lynnie Trzoss / Davide Povero / Milos Lazic / Geza Ambrus-Aikelin / Angelina Santini / Rama Pranadinata / Gretchen Bain / Ryan Stansfield / Jeffrey A Stafford / James Veal / Ryan Takahashi / Justin Ly / Shu Chen / Liling Liu / Marika Nespi / Robert Blake / Arna Katewa / Tracy Kleinheinz / Swathi Sujatha-Bhaskar / Nandhini Ramamoorthi / Jessica Sims / Brent McKenzie / Mark Chen / Mark Ultsch / Matthew Johnson / Jeremy Murray / Claudio Ciferri / Steven T Staben / Michael J Townsend / Craig E Stivala / Abstract: Inappropriate activation of the NLRP3 inflammasome has been implicated in multiple inflammatory and autoimmune diseases. Herein, we aimed to develop novel NLRP3 inhibitors that could minimize the ...Inappropriate activation of the NLRP3 inflammasome has been implicated in multiple inflammatory and autoimmune diseases. Herein, we aimed to develop novel NLRP3 inhibitors that could minimize the risk of drug-induced liver injury. Lipophilic ligand efficiency was used as a guiding metric to identify a series of 6,7-dihydro-5H-pyrazolo[5,1-][1,3]oxazinesulfonylureas. A leading compound from this series was advanced into safety studies in cynomolgus monkeys, and renal toxicity, due to compound precipitation, was observed. To overcome this obstacle, we focused on improving the solubility of our compounds, specifically by introducing basic amine substituents into the scaffold. This led to the identification of GDC-2394, a potent and selective NLRP3 inhibitor, with an in vitro and in vivo safety profile suitable for advancement into human clinical trials. | |||||||||
History |
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-Structure visualization
Supplemental images |
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-Downloads & links
-EMDB archive
Map data | emd_28596.map.gz | 2.2 MB | EMDB map data format | |
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Header (meta data) | emd-28596-v30.xml emd-28596.xml | 21.3 KB 21.3 KB | Display Display | EMDB header |
FSC (resolution estimation) | emd_28596_fsc.xml | 10.6 KB | Display | FSC data file |
Images | emd_28596.png | 179.7 KB | ||
Filedesc metadata | emd-28596.cif.gz | 6.9 KB | ||
Others | emd_28596_half_map_1.map.gz emd_28596_half_map_2.map.gz | 8 MB 8 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-28596 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-28596 | HTTPS FTP |
-Related structure data
Related structure data | 8etrMC M: atomic model generated by this map C: citing same article (ref.) |
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Similar structure data | Similarity search - Function & homologyF&H Search |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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Related items in Molecule of the Month |
-Map
File | Download / File: emd_28596.map.gz / Format: CCP4 / Size: 2.3 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||
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Voxel size | X=Y=Z: 1.1093 Å | ||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||
Details | EMDB XML:
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-Supplemental data
-Half map: #1
File | emd_28596_half_map_1.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Half map: #2
File | emd_28596_half_map_2.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Sample components
-Entire : NLRP3 NACHT domain
Entire | Name: NLRP3 NACHT domain |
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Components |
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-Supramolecule #1: NLRP3 NACHT domain
Supramolecule | Name: NLRP3 NACHT domain / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1 / Details: NACHT domain only |
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Source (natural) | Organism: Homo sapiens (human) |
Molecular weight | Theoretical: 120 KDa |
-Macromolecule #1: NACHT, LRR and PYD domains-containing protein 3
Macromolecule | Name: NACHT, LRR and PYD domains-containing protein 3 / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: Homo sapiens (human) |
Molecular weight | Theoretical: 63.38034 KDa |
Recombinant expression | Organism: Spodoptera frugiperda (fall armyworm) |
Sequence | String: KDYRKKYRKY VRSRFQCIED RNARLGESVS LNKRYTRLRL IKEHRSQQER EQELLAIGKT KTCESPVSPI KMELLFDPDD EHSEPVHTV VFQGAAGIGK TILARKMMLD WASGTLYQDR FDYLFYIHCR EVSLVTQRSL GDLIMSCCPD PNPPIHKIVR K PSRILFLM ...String: KDYRKKYRKY VRSRFQCIED RNARLGESVS LNKRYTRLRL IKEHRSQQER EQELLAIGKT KTCESPVSPI KMELLFDPDD EHSEPVHTV VFQGAAGIGK TILARKMMLD WASGTLYQDR FDYLFYIHCR EVSLVTQRSL GDLIMSCCPD PNPPIHKIVR K PSRILFLM DGFDELQGAF DEHIGPLCTD WQKAERGDIL LSSLIRKKLL PEASLLITTR PVALEKLQHL LDHPRHVEIL GF SEAKRKE YFFKYFSDEA QARAAFSLIQ ENEVLFTMCF IPLVCWIVCT GLKQQMESGK SLAQTSKTTT AVYVFFLSSL LQP RGGSQE HGLCAHLWGL CSLAADGIWN QKILFEESDL RNHGLQKADV SAFLRMNLFQ KEVDCEKFYS FIHMTFQEFF AAMY YLLEE EKEGRTNVPG SRLKLPSRDV TVLLENYGKF EKGYLIFVVR FLFGLVNQER TSYLEKKLSC KISQQIRLEL LKWIE VKAK AKKLQIQPSQ LELFYCLYEM QEEDFVQRAM DYFPKIEINL STRMDHMVSS FCIENCHRV UniProtKB: NACHT, LRR and PYD domains-containing protein 3 |
-Macromolecule #2: MAGNESIUM ION
Macromolecule | Name: MAGNESIUM ION / type: ligand / ID: 2 / Number of copies: 1 / Formula: MG |
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Molecular weight | Theoretical: 24.305 Da |
-Macromolecule #3: ADENOSINE-5'-DIPHOSPHATE
Macromolecule | Name: ADENOSINE-5'-DIPHOSPHATE / type: ligand / ID: 3 / Number of copies: 1 / Formula: ADP |
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Molecular weight | Theoretical: 427.201 Da |
Chemical component information | ChemComp-ADP: |
-Macromolecule #4: (6S,8R)-N-[(1,2,3,5,6,7-hexahydro-s-indacen-4-yl)carbamoyl]-6-(me...
Macromolecule | Name: (6S,8R)-N-[(1,2,3,5,6,7-hexahydro-s-indacen-4-yl)carbamoyl]-6-(methylamino)-6,7-dihydro-5H-pyrazolo[5,1-b][1,3]oxazine-3-sulfonamide type: ligand / ID: 4 / Number of copies: 1 / Formula: WTN |
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Molecular weight | Theoretical: 431.509 Da |
Chemical component information | ChemComp-WTN: |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 0.5 mg/mL | |||||||||||||||
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Buffer | pH: 7.5 Component:
Details: 0.15M NaCl, 20mM Tris pH 7.5, 10% glycerol, 1mM TCEP, 2.5mM ATP, 2mM MgCl2. | |||||||||||||||
Grid | Model: Quantifoil R1.2/1.3 / Material: GOLD / Support film - Material: GOLD / Support film - topology: HOLEY / Support film - Film thickness: 25 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 20 sec. / Pretreatment - Pressure: 0.007 kPa Details: Holey gold grids (UltrAuFoil 25 nm R 1.2/1.3) were glow discharged for 20 seconds using a Solarus plasma cleaner (Gatan, Pleasanton, CA, USA), and grids were blotted and plunge frozen in ...Details: Holey gold grids (UltrAuFoil 25 nm R 1.2/1.3) were glow discharged for 20 seconds using a Solarus plasma cleaner (Gatan, Pleasanton, CA, USA), and grids were blotted and plunge frozen in liquid ethane using a Vitrobot (ThermoFisher Scientific, Waltham, MA), operating at 4C, 100 relative humidity, blot force 7, and with a 4 second blot time. | |||||||||||||||
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK III | |||||||||||||||
Details | mono disperse |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Software | Name: SerialEM |
Image recording | Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Average exposure time: 3.0 sec. / Average electron dose: 64.0 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | C2 aperture diameter: 50.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 3.0 µm / Nominal defocus min: 0.6 µm |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
+Image processing
-Atomic model buiding 1
Software | Name: Coot |
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Refinement | Space: REAL / Protocol: OTHER |
Output model | PDB-8etr: |