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Yorodumi- EMDB-7831: Cryo-EM structure of FLNaABD E254K bound to phalloidin-stabilized... -
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-Basic information
Entry | Database: EMDB / ID: EMD-7831 | |||||||||
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Title | Cryo-EM structure of FLNaABD E254K bound to phalloidin-stabilized F-actin | |||||||||
Map data | Symmetrized map of FLNaABD-E254K bound to phalloidin-stabilized F-actin. This map has been low-pass filtered to 3.6 A and sharpened with a B-factor of -150. | |||||||||
Sample |
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Keywords | Actin-binding domain / Actin crosslinker / STRUCTURAL PROTEIN | |||||||||
Function / homology | Function and homology information regulation of membrane repolarization during atrial cardiac muscle cell action potential / regulation of membrane repolarization during cardiac muscle cell action potential / establishment of Sertoli cell barrier / Myb complex / formation of radial glial scaffolds / glycoprotein Ib-IX-V complex / adenylate cyclase-inhibiting dopamine receptor signaling pathway / positive regulation of integrin-mediated signaling pathway / cytoplasmic sequestering of protein / tubulin deacetylation ...regulation of membrane repolarization during atrial cardiac muscle cell action potential / regulation of membrane repolarization during cardiac muscle cell action potential / establishment of Sertoli cell barrier / Myb complex / formation of radial glial scaffolds / glycoprotein Ib-IX-V complex / adenylate cyclase-inhibiting dopamine receptor signaling pathway / positive regulation of integrin-mediated signaling pathway / cytoplasmic sequestering of protein / tubulin deacetylation / actin crosslink formation / OAS antiviral response / blood coagulation, intrinsic pathway / protein localization to bicellular tight junction / Striated Muscle Contraction / positive regulation of actin filament bundle assembly / positive regulation of neuron migration / Cell-extracellular matrix interactions / positive regulation of potassium ion transmembrane transport / early endosome to late endosome transport / apical dendrite / cell-cell junction organization / positive regulation of neural precursor cell proliferation / positive regulation of platelet activation / protein localization to cell surface / negative regulation of transcription by RNA polymerase I / Fc-gamma receptor I complex binding / wound healing, spreading of cells / megakaryocyte development / GP1b-IX-V activation signalling / cortical cytoskeleton / receptor clustering / positive regulation of axon regeneration / actin filament bundle / SMAD binding / RHO GTPases activate PAKs / skeletal muscle thin filament assembly / brush border / striated muscle thin filament / semaphorin-plexin signaling pathway / cilium assembly / mitotic spindle assembly / potassium channel regulator activity / epithelial to mesenchymal transition / blood vessel remodeling / skeletal muscle fiber development / axonal growth cone / heart morphogenesis / positive regulation of substrate adhesion-dependent cell spreading / stress fiber / regulation of cell migration / release of sequestered calcium ion into cytosol / protein kinase C binding / dendritic shaft / G protein-coupled receptor binding / actin filament / protein localization to plasma membrane / synapse organization / Hydrolases; Acting on acid anhydrides; Acting on acid anhydrides to facilitate cellular and subcellular movement / mRNA transcription by RNA polymerase II / establishment of protein localization / trans-Golgi network / negative regulation of DNA-binding transcription factor activity / negative regulation of protein catabolic process / cerebral cortex development / platelet aggregation / small GTPase binding / Z disc / kinase binding / positive regulation of protein import into nucleus / actin filament binding / cell-cell junction / actin cytoskeleton / negative regulation of neuron projection development / Platelet degranulation / GTPase binding / toxin activity / perikaryon / actin cytoskeleton organization / postsynapse / angiogenesis / positive regulation of canonical NF-kappaB signal transduction / DNA-binding transcription factor binding / transmembrane transporter binding / protein stabilization / hydrolase activity / cadherin binding / focal adhesion / glutamatergic synapse / nucleolus / negative regulation of apoptotic process / perinuclear region of cytoplasm / protein homodimerization activity / RNA binding / extracellular exosome / extracellular region / ATP binding / membrane / nucleus / plasma membrane Similarity search - Function | |||||||||
Biological species | Homo sapiens (human) / Gallus gallus (chicken) / Amanita phalloides (death cap) | |||||||||
Method | helical reconstruction / cryo EM / Resolution: 3.54 Å | |||||||||
Authors | Iwamoto DV / Huehn AR | |||||||||
Citation | Journal: Nat Struct Mol Biol / Year: 2018 Title: Structural basis of the filamin A actin-binding domain interaction with F-actin. Authors: Daniel V Iwamoto / Andrew Huehn / Bertrand Simon / Clotilde Huet-Calderwood / Massimiliano Baldassarre / Charles V Sindelar / David A Calderwood / Abstract: Actin-cross-linking proteins assemble actin filaments into higher-order structures essential for orchestrating cell shape, adhesion, and motility. Missense mutations in the tandem calponin homology ...Actin-cross-linking proteins assemble actin filaments into higher-order structures essential for orchestrating cell shape, adhesion, and motility. Missense mutations in the tandem calponin homology domains of their actin-binding domains (ABDs) underlie numerous genetic diseases, but a molecular understanding of these pathologies is hampered by the lack of high-resolution structures of any actin-cross-linking protein bound to F-actin. Here, taking advantage of a high-affinity, disease-associated mutant of the human filamin A (FLNa) ABD, we combine cryo-electron microscopy and functional studies to reveal at near-atomic resolution how the first calponin homology domain (CH1) and residues immediately N-terminal to it engage actin. We further show that reorientation of CH2 relative to CH1 is required to avoid clashes with actin and to expose F-actin-binding residues on CH1. Our data explain localization of disease-associated loss-of-function mutations to FLNaCH1 and gain-of-function mutations to the regulatory FLNaCH2. Sequence conservation argues that this provides a general model for ABD-F-actin binding. | |||||||||
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_7831.map.gz | 5.4 MB | EMDB map data format | |
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Header (meta data) | emd-7831-v30.xml emd-7831.xml | 22.3 KB 22.3 KB | Display Display | EMDB header |
FSC (resolution estimation) | emd_7831_fsc.xml | 7.4 KB | Display | FSC data file |
Images | emd_7831.png | 74 KB | ||
Masks | emd_7831_msk_1.map | 35.3 MB | Mask map | |
Filedesc metadata | emd-7831.cif.gz | 6.9 KB | ||
Others | emd_7831_additional.map.gz emd_7831_half_map_1.map.gz emd_7831_half_map_2.map.gz | 7.4 MB 11.9 MB 11.9 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-7831 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-7831 | HTTPS FTP |
-Related structure data
Related structure data | 6d8cMC 7832C 7833C 8918C C: citing same article (ref.) M: atomic model generated by this map |
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Similar structure data |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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Related items in Molecule of the Month |
-Map
File | Download / File: emd_7831.map.gz / Format: CCP4 / Size: 35.3 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | Symmetrized map of FLNaABD-E254K bound to phalloidin-stabilized F-actin. This map has been low-pass filtered to 3.6 A and sharpened with a B-factor of -150. | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 1.33 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_7831_msk_1.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Additional map: Symmetrized map of FLNaABD-E254K bound to phalloidin-stabilized F-actin....
File | emd_7831_additional.map | ||||||||||||
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Annotation | Symmetrized map of FLNaABD-E254K bound to phalloidin-stabilized F-actin. Low-pass filtered to 10 A and sharpened with a B-factor of -150. Masked to display the FLNaABD CH2 domain cryo-EM density. | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: Independently refined half map(2) of FLNaABD-E254K bound to...
File | emd_7831_half_map_1.map | ||||||||||||
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Annotation | Independently refined half map(2) of FLNaABD-E254K bound to phalloidin-stabilized F-actin. This half map was symmetrized using parameters calculated from the full map. | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: Independently refined half map(1) of FLNaABD-E254K bound to...
File | emd_7831_half_map_2.map | ||||||||||||
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Annotation | Independently refined half map(1) of FLNaABD-E254K bound to phalloidin-stabilized F-actin. This half map was symmetrized using parameters calculated from the full map. | ||||||||||||
Projections & Slices |
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Density Histograms |
-Sample components
-Entire : Helical complex of FLNaABD E254K bound to phalloidin-stabilized F...
Entire | Name: Helical complex of FLNaABD E254K bound to phalloidin-stabilized F-actin |
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Components |
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-Supramolecule #1: Helical complex of FLNaABD E254K bound to phalloidin-stabilized F...
Supramolecule | Name: Helical complex of FLNaABD E254K bound to phalloidin-stabilized F-actin type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#3 |
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Source (natural) | Organism: Homo sapiens (human) |
-Macromolecule #1: Filamin-A
Macromolecule | Name: Filamin-A / type: protein_or_peptide / ID: 1 / Number of copies: 5 / Enantiomer: LEVO |
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Source (natural) | Organism: Homo sapiens (human) |
Molecular weight | Theoretical: 34.47625 KDa |
Recombinant expression | Organism: Escherichia coli (E. coli) |
Sequence | String: MHHHHHHGSL VPRSENLYFQ GSDILQGTMS SSHSRAGQSA AGAAPGGGVD TRDAEMPATE KDLAEDAPWK KIQQNTFTRW CNEHLKCVS KRIANLQTDL SDGLRLIALL EVLSQKKMHR KHNQRPTFRQ MQLENVSVAL EFLDRESIKL VSIDSKAIVD G NLKLILGL ...String: MHHHHHHGSL VPRSENLYFQ GSDILQGTMS SSHSRAGQSA AGAAPGGGVD TRDAEMPATE KDLAEDAPWK KIQQNTFTRW CNEHLKCVS KRIANLQTDL SDGLRLIALL EVLSQKKMHR KHNQRPTFRQ MQLENVSVAL EFLDRESIKL VSIDSKAIVD G NLKLILGL IWTLILHYSI SMPMWDEEED EEAKKQTPKQ RLLGWIQNKL PQLPITNFSR DWQSGRALGA LVDSCAPGLC PD WDSWDAS KPVTNAREAM QQADDWLGIP QVITPEEIVD PNVDKHSVMT YLSQFPKAKL KPGAPLRPK UniProtKB: Filamin-A |
-Macromolecule #2: Actin, alpha skeletal muscle
Macromolecule | Name: Actin, alpha skeletal muscle / type: protein_or_peptide / ID: 2 / Number of copies: 5 / Enantiomer: LEVO |
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Source (natural) | Organism: Gallus gallus (chicken) / Tissue: Breast |
Molecular weight | Theoretical: 41.862613 KDa |
Sequence | String: DEDETTALVC DNGSGLVKAG FAGDDAPRAV FPSIVGRPRH QGVMVGMGQK DSYVGDEAQS KRGILTLKYP IEHGIITNWD DMEKIWHHT FYNELRVAPE EHPTLLTEAP LNPKANREKM TQIMFETFNV PAMYVAIQAV LSLYASGRTT GIVLDSGDGV T HNVPIYEG ...String: DEDETTALVC DNGSGLVKAG FAGDDAPRAV FPSIVGRPRH QGVMVGMGQK DSYVGDEAQS KRGILTLKYP IEHGIITNWD DMEKIWHHT FYNELRVAPE EHPTLLTEAP LNPKANREKM TQIMFETFNV PAMYVAIQAV LSLYASGRTT GIVLDSGDGV T HNVPIYEG YALPHAIMRL DLAGRDLTDY LMKILTERGY SFVTTAEREI VRDIKEKLCY VALDFENEMA TAASSSSLEK SY ELPDGQV ITIGNERFRC PETLFQPSFI GMESAGIHET TYNSIMKCDI DIRKDLYANN VMSGGTTMYP GIADRMQKEI TAL APSTMK IKIIAPPERK YSVWIGGSIL ASLSTFQQMW ITKQEYDEAG PSIVHRKCF UniProtKB: Actin, alpha skeletal muscle |
-Macromolecule #3: Phalloidin
Macromolecule | Name: Phalloidin / type: protein_or_peptide / ID: 3 / Number of copies: 3 / Enantiomer: LEVO |
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Source (natural) | Organism: Amanita phalloides (death cap) |
Molecular weight | Theoretical: 808.899 Da |
Sequence | String: (HYP)AW(G5G)A(ALO)C |
-Macromolecule #4: ADENOSINE-5'-DIPHOSPHATE
Macromolecule | Name: ADENOSINE-5'-DIPHOSPHATE / type: ligand / ID: 4 / Number of copies: 5 / Formula: ADP |
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Molecular weight | Theoretical: 427.201 Da |
Chemical component information | ChemComp-ADP: |
-Macromolecule #5: MAGNESIUM ION
Macromolecule | Name: MAGNESIUM ION / type: ligand / ID: 5 / Number of copies: 5 / Formula: MG |
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Molecular weight | Theoretical: 24.305 Da |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | helical reconstruction |
Aggregation state | filament |
-Sample preparation
Buffer | pH: 8 |
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Vitrification | Cryogen name: ETHANE / Instrument: HOMEMADE PLUNGER |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | C2 aperture diameter: 50.0 µm / Calibrated magnification: 37500 / Illumination mode: SPOT SCAN / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 2.9 µm / Nominal defocus min: 1.0 µm / Nominal magnification: 105000 |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Image recording | Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: SUPER-RESOLUTION / Digitization - Frames/image: 1-40 / Number grids imaged: 2 / Number real images: 2140 / Average exposure time: 12.0 sec. / Average electron dose: 50.0 e/Å2 Details: Micrographs from only one of the grids were used in the reconstruction. From that grid, only micrographs where Gctf detected signal at resolutions better than 4A were used in the reconstruction (~15%). |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
-Image processing
-Atomic model buiding 1
Initial model |
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Refinement | Protocol: RIGID BODY FIT | ||||||
Output model | PDB-6d8c: |