+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-40302 | |||||||||
---|---|---|---|---|---|---|---|---|---|---|
Title | The consensus map of a 12-LOX hexamer | |||||||||
Map data | The consensus map of a 12-LOX hexamer | |||||||||
Sample |
| |||||||||
Keywords | Lipoxygenase / platelets / lipid-modifying enzyme / lipid oxidation / OXIDOREDUCTASE | |||||||||
Biological species | Homo sapiens (human) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 2.63 Å | |||||||||
Authors | Black KA / Mobbs JI / Venugopal H / Thal DM / Glukhova A | |||||||||
Funding support | United States, 2 items
| |||||||||
Citation | Journal: Blood / Year: 2023 Title: Cryo-EM structures of human arachidonate 12S-lipoxygenase bound to endogenous and exogenous inhibitors. Authors: Jesse I Mobbs / Katrina A Black / Michelle Tran / Wessel A C Burger / Hariprasad Venugopal / Theodore R Holman / Michael Holinstat / David M Thal / Alisa Glukhova / Abstract: Human 12-lipoxygenase (12-LOX) is a key enzyme involved in platelet activation, and the regulation of its activity has been targeted for the treatment of heparin-induced thrombocytopenia. Despite the ...Human 12-lipoxygenase (12-LOX) is a key enzyme involved in platelet activation, and the regulation of its activity has been targeted for the treatment of heparin-induced thrombocytopenia. Despite the clinical importance of 12-LOX, the exact mechanisms by which it affects platelet activation are not fully understood, and the lack of structural information has limited drug discovery efforts. In this study, we used single-particle cryo-electron microscopy to determine high-resolution structures (1.7-2.8 Å) of human 12-LOX. Our results showed that 12-LOX can exist in multiple oligomeric states, from monomer to hexamer, which may affect its catalytic activity and membrane association. We also identified different conformations within the 12-LOX dimer, which likely represent different time points in its catalytic cycle. Furthermore, we identified small molecules bound to 12-LOX. The active site of the 12-LOX tetramer was occupied by an endogenous 12-LOX inhibitor, a long-chain acyl coenzyme A. In addition, we found that the 12-LOX hexamer can simultaneously bind to arachidonic acid and ML355, a selective 12-LOX inhibitor that has passed a phase 1 clinical trial for the treatment of heparin-induced thrombocytopenia and received a fast-track designation by the Food and Drug Administration. Overall, our findings provide novel insights into the assembly of 12-LOX oligomers, their catalytic mechanism, and small molecule binding, paving the way for further drug development targeting the 12-LOX enzyme. | |||||||||
History |
|
-Structure visualization
Supplemental images |
---|
-Downloads & links
-EMDB archive
Map data | emd_40302.map.gz | 62.6 MB | EMDB map data format | |
---|---|---|---|---|
Header (meta data) | emd-40302-v30.xml emd-40302.xml | 14.2 KB 14.2 KB | Display Display | EMDB header |
FSC (resolution estimation) | emd_40302_fsc.xml | 11.4 KB | Display | FSC data file |
Images | emd_40302.png | 172.8 KB | ||
Masks | emd_40302_msk_1.map | 125 MB | Mask map | |
Filedesc metadata | emd-40302.cif.gz | 5.1 KB | ||
Others | emd_40302_half_map_1.map.gz emd_40302_half_map_2.map.gz | 115.9 MB 115.9 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-40302 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-40302 | HTTPS FTP |
-Validation report
Summary document | emd_40302_validation.pdf.gz | 1 MB | Display | EMDB validaton report |
---|---|---|---|---|
Full document | emd_40302_full_validation.pdf.gz | 1 MB | Display | |
Data in XML | emd_40302_validation.xml.gz | 19 KB | Display | |
Data in CIF | emd_40302_validation.cif.gz | 24.9 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-40302 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-40302 | HTTPS FTP |
-Related structure data
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
---|
-Map
File | Download / File: emd_40302.map.gz / Format: CCP4 / Size: 125 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Annotation | The consensus map of a 12-LOX hexamer | ||||||||||||||||||||
Voxel size | X=Y=Z: 0.82 Å | ||||||||||||||||||||
Density |
| ||||||||||||||||||||
Symmetry | Space group: 1 | ||||||||||||||||||||
Details | EMDB XML:
|
-Supplemental data
-Mask #1
File | emd_40302_msk_1.map | ||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Projections & Slices |
| ||||||||||||
Density Histograms |
-Half map: Half map 1 from a non-uniform refinement a 12-LOX hexamer
File | emd_40302_half_map_1.map | ||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Annotation | Half map 1 from a non-uniform refinement a 12-LOX hexamer | ||||||||||||
Projections & Slices |
| ||||||||||||
Density Histograms |
-Half map: Half map 2 from a non-uniform refinement a 12-LOX hexamer
File | emd_40302_half_map_2.map | ||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Annotation | Half map 2 from a non-uniform refinement a 12-LOX hexamer | ||||||||||||
Projections & Slices |
| ||||||||||||
Density Histograms |
-Sample components
-Entire : Dimeric human 12-Lipoxygenase
Entire | Name: Dimeric human 12-Lipoxygenase |
---|---|
Components |
|
-Supramolecule #1: Dimeric human 12-Lipoxygenase
Supramolecule | Name: Dimeric human 12-Lipoxygenase / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all |
---|---|
Source (natural) | Organism: Homo sapiens (human) |
-Macromolecule #1: Polyunsaturated fatty acid lipoxygenase ALOX12
Macromolecule | Name: Polyunsaturated fatty acid lipoxygenase ALOX12 / type: protein_or_peptide / ID: 1 / Enantiomer: LEVO |
---|---|
Source (natural) | Organism: Homo sapiens (human) |
Sequence | String: MHHHHHHGRY RIRVATGAWL FSGSYNRVQL WLVGTRGEAE LELQLRPARG EEEEFDHDVA EDLGLLQFVR LRKHHWLVDD AWFCDRITV QGPGACAEVA FPCYRWVQGE DILSLPEGTA RLPGDNALDM FQKHREKELK DRQQIYCWAT WKEGLPLTIA A DRKDDLPP ...String: MHHHHHHGRY RIRVATGAWL FSGSYNRVQL WLVGTRGEAE LELQLRPARG EEEEFDHDVA EDLGLLQFVR LRKHHWLVDD AWFCDRITV QGPGACAEVA FPCYRWVQGE DILSLPEGTA RLPGDNALDM FQKHREKELK DRQQIYCWAT WKEGLPLTIA A DRKDDLPP NMRFHEEKRL DFEWTLKAGA LEMALKRVYT LLSSWNCLED FDQIFWGQKS ALAEKVRQCW QDDELFSYQF LN GANPMLL RRSTSLPSRL VLPSGMEELQ AQLEKELQNG SLFEADFILL DGIPANVIRG EKQYLAAPLV MLKMEPNGKL QPM VIQIQP PSPSSPTPTL FLPSDPPLAW LLAKSWVRNS DFQLHEIQYH LLNTHLVAEV IAVATMRCLP GLHPIFKFLI PHIR YTMEI NTRARTQLIS DGGIFDKAVS TGGGGHVQLL RRAAAQLTYC SLCPPDDLAD RGLLGLPGAL YAHDALRLWE IIARY VEGI VHLFYQRDDI VKGDPELQAW CREITEVGLC QAQDRGFPVS FQSQSQLCHF LTMCVFTCTA QHAAINQGQL DWYAWV PNA PCTMRMPPPT TKEDVTMATV MGSLPDVRQA CLQMAISWHL SRRQPDMVPL GHHKEKYFSG PKPKAVLNQF RTDLEKL EK EITARNEQLD WPYEYLKPSC IENSVTI |
-Experimental details
-Structure determination
Method | cryo EM |
---|---|
Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Buffer | pH: 8 |
---|---|
Vitrification | Cryogen name: ETHANE |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
---|---|
Image recording | Film or detector model: GATAN K3 (6k x 4k) / Average electron dose: 60.0 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | C2 aperture diameter: 50.0 µm / Calibrated magnification: 105000 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 1.5 µm / Nominal defocus min: 0.5 µm |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
+Image processing
-Atomic model buiding 1
Refinement | Protocol: FLEXIBLE FIT |
---|