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- EMDB-8886: Cryo-EM structure of F-actin complexed with the beta-III-spectrin... -
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Open data
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Basic information
Entry | Database: EMDB / ID: EMD-8886 | ||||||||||||
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Title | Cryo-EM structure of F-actin complexed with the beta-III-spectrin actin-binding domain | ||||||||||||
![]() | Cryo-EM structure of F-actin complexed with the beta-III-spectrin actin-binding domain | ||||||||||||
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![]() | actin binding protein / filament / STRUCTURAL PROTEIN | ||||||||||||
Function / homology | ![]() cerebellar Purkinje cell layer morphogenesis / structural constituent of synapse / postsynaptic spectrin-associated cytoskeleton / structural constituent of postsynapse / regulation of postsynaptic specialization assembly / spectrin / positive regulation of norepinephrine uptake / cellular response to cytochalasin B / regulation of transepithelial transport / paranodal junction ...cerebellar Purkinje cell layer morphogenesis / structural constituent of synapse / postsynaptic spectrin-associated cytoskeleton / structural constituent of postsynapse / regulation of postsynaptic specialization assembly / spectrin / positive regulation of norepinephrine uptake / cellular response to cytochalasin B / regulation of transepithelial transport / paranodal junction / morphogenesis of a polarized epithelium / bBAF complex / postsynaptic actin cytoskeleton organization / npBAF complex / nBAF complex / protein localization to adherens junction / brahma complex / postsynaptic actin cytoskeleton / Tat protein binding / structural constituent of postsynaptic actin cytoskeleton / GBAF complex / dense body / Formation of annular gap junctions / regulation of G0 to G1 transition / Gap junction degradation / Cell-extracellular matrix interactions / Folding of actin by CCT/TriC / Regulation of MITF-M-dependent genes involved in pigmentation / apical protein localization / regulation of double-strand break repair / actin filament capping / adherens junction assembly / regulation of nucleotide-excision repair / Prefoldin mediated transfer of substrate to CCT/TriC / RSC-type complex / RHOF GTPase cycle / Adherens junctions interactions / tight junction / Sensory processing of sound by outer hair cells of the cochlea / regulation of norepinephrine uptake / Interaction between L1 and Ankyrins / Sensory processing of sound by inner hair cells of the cochlea / regulation of mitotic metaphase/anaphase transition / SWI/SNF complex / parallel fiber to Purkinje cell synapse / positive regulation of double-strand break repair / regulation of synaptic vesicle endocytosis / positive regulation of T cell differentiation / apical junction complex / establishment or maintenance of cell polarity / adult behavior / cortical actin cytoskeleton / regulation of cyclin-dependent protein serine/threonine kinase activity / cortical cytoskeleton / maintenance of blood-brain barrier / positive regulation of stem cell population maintenance / NuA4 histone acetyltransferase complex / nitric-oxide synthase binding / regulation of G1/S transition of mitotic cell cycle / Recycling pathway of L1 / kinesin binding / brush border / calyx of Held / negative regulation of cell differentiation / positive regulation of double-strand break repair via homologous recombination / EPH-ephrin mediated repulsion of cells / RHO GTPases Activate WASPs and WAVEs / RHO GTPases activate IQGAPs / regulation of protein localization to plasma membrane / positive regulation of myoblast differentiation / COPI-mediated anterograde transport / vesicle-mediated transport / synapse assembly / EPHB-mediated forward signaling / substantia nigra development / MHC class II antigen presentation / NCAM signaling for neurite out-growth / axonogenesis / negative regulation of protein binding / Translocation of SLC2A4 (GLUT4) to the plasma membrane / cell projection / cell motility / actin filament / RHO GTPases Activate Formins / positive regulation of cell differentiation / adherens junction / FCGR3A-mediated phagocytosis / regulation of transmembrane transporter activity / Hydrolases; Acting on acid anhydrides; Acting on acid anhydrides to facilitate cellular and subcellular movement / Signaling by high-kinase activity BRAF mutants / DNA Damage Recognition in GG-NER / MAP2K and MAPK activation / tau protein binding / Schaffer collateral - CA1 synapse / multicellular organism growth / B-WICH complex positively regulates rRNA expression / structural constituent of cytoskeleton / phospholipid binding / kinetochore / Regulation of actin dynamics for phagocytic cup formation Similarity search - Function | ||||||||||||
Biological species | ![]() | ||||||||||||
Method | helical reconstruction / cryo EM / negative staining / Resolution: 7.0 Å | ||||||||||||
![]() | Wang F / Orlova A | ||||||||||||
Funding support | ![]()
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![]() | ![]() Title: Structural basis for high-affinity actin binding revealed by a β-III-spectrin SCA5 missense mutation. Authors: Adam W Avery / Michael E Fealey / Fengbin Wang / Albina Orlova / Andrew R Thompson / David D Thomas / Thomas S Hays / Edward H Egelman / ![]() Abstract: Spinocerebellar ataxia type 5 (SCA5) is a neurodegenerative disease caused by mutations in the cytoskeletal protein β-III-spectrin. Previously, a SCA5 mutation resulting in a leucine-to-proline ...Spinocerebellar ataxia type 5 (SCA5) is a neurodegenerative disease caused by mutations in the cytoskeletal protein β-III-spectrin. Previously, a SCA5 mutation resulting in a leucine-to-proline substitution (L253P) in the actin-binding domain (ABD) was shown to cause a 1000-fold increase in actin-binding affinity. However, the structural basis for this increase is unknown. Here, we report a 6.9 Å cryo-EM structure of F-actin complexed with the L253P ABD. This structure, along with co-sedimentation and pulsed-EPR measurements, demonstrates that high-affinity binding caused by the CH2-localized mutation is due to opening of the two CH domains. This enables CH1 to bind actin aided by an unstructured N-terminal region that becomes α-helical upon binding. This helix is required for association with actin as truncation eliminates binding. Collectively, these results shed light on the mechanism by which β-III-spectrin, and likely similar actin-binding proteins, interact with actin, and how this mechanism can be perturbed to cause disease. | ||||||||||||
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 | ![]() | 24.6 MB | ![]() | |
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Header (meta data) | ![]() ![]() | 12.7 KB 12.7 KB | Display Display | ![]() |
Images | ![]() | 180.3 KB | ||
Filedesc metadata | ![]() | 5.8 KB | ||
Archive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 6anuMC 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 structure of F-actin complexed with the beta-III-spectrin actin-binding domain | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 1.05 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
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Sample components
-Entire : F-actin complexed with the spectrin actin-binding domain
Entire | Name: F-actin complexed with the spectrin actin-binding domain |
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Components |
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-Supramolecule #1: F-actin complexed with the spectrin actin-binding domain
Supramolecule | Name: F-actin complexed with the spectrin actin-binding domain type: complex / ID: 1 / Parent: 0 / Macromolecule list: all |
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Source (natural) | Organism: ![]() |
-Macromolecule #1: Actin, cytoplasmic 1
Macromolecule | Name: Actin, cytoplasmic 1 / type: protein_or_peptide / ID: 1 / Number of copies: 6 / Enantiomer: LEVO |
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Source (natural) | Organism: ![]() |
Molecular weight | Theoretical: 41.78266 KDa |
Recombinant expression | Organism: ![]() ![]() |
Sequence | String: MDDDIAALVV DNGSGMCKAG FAGDDAPRAV FPSIVGRPRH QGVMVGMGQK DSYVGDEAQS KRGILTLKYP IEHGIVTNWD DMEKIWHHT FYNELRVAPE EHPVLLTEAP LNPKANREKM TQIMFETFNT PAMYVAIQAV LSLYASGRTT GIVMDSGDGV T HTVPIYEG ...String: MDDDIAALVV DNGSGMCKAG FAGDDAPRAV FPSIVGRPRH QGVMVGMGQK DSYVGDEAQS KRGILTLKYP IEHGIVTNWD DMEKIWHHT FYNELRVAPE EHPVLLTEAP LNPKANREKM TQIMFETFNT PAMYVAIQAV LSLYASGRTT GIVMDSGDGV T HTVPIYEG YALPHAILRL DLAGRDLTDY LMKILTERGY SFTTTAEREI VRDIKEKLCY VALDFEQEMA TAASSSSLEK SY ELPDGQV ITIGNERFRC PEALFQPSFL GMESCGIHET TFNSIMKCDV DIRKDLYANT VLSGGTTMYP GIADRMQKEI TAL APSTMK IKIIAPPERK YSVWIGGSIL ASLSTFQQMW ISKQEYDESG PSIVHRKCF UniProtKB: Actin, cytoplasmic 1 |
-Macromolecule #2: Spectrin beta chain, non-erythrocytic 2
Macromolecule | Name: Spectrin beta chain, non-erythrocytic 2 / type: protein_or_peptide / ID: 2 / Number of copies: 6 / Enantiomer: LEVO |
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Source (natural) | Organism: ![]() |
Molecular weight | Theoretical: 32.857141 KDa |
Recombinant expression | Organism: ![]() ![]() |
Sequence | String: MSSTLSPTDF DSLEIQGQYS DINNRWDLPD SDWDNDSSSA RLFERSRIKA LADEREAVQK KTFTKWVNSH LARVTCRVGD LYSDLRDGR NLLRLLEVLS GEILPKPTKG RMRIHCLENV DKALQFLKEQ KVHLENMGSH DIVDGNHRLT LGLVWTIILR F QIQDISVE ...String: MSSTLSPTDF DSLEIQGQYS DINNRWDLPD SDWDNDSSSA RLFERSRIKA LADEREAVQK KTFTKWVNSH LARVTCRVGD LYSDLRDGR NLLRLLEVLS GEILPKPTKG RMRIHCLENV DKALQFLKEQ KVHLENMGSH DIVDGNHRLT LGLVWTIILR F QIQDISVE TEDNKEKKSA KDALLLWCQM KTAGYPNVNV HNFTTSWRDG LAFNAIVHKH RPDLLDFESL KKCNAHYNLQ NA FNLAEKE LGLTKPLDPE DVNVDQPDEK SIITYVATYY HYFSKMK UniProtKB: Spectrin beta chain, non-erythrocytic 2 |
-Experimental details
-Structure determination
Method | negative staining, cryo EM |
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![]() | helical reconstruction |
Aggregation state | filament |
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Sample preparation
Buffer | pH: 7.4 |
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Staining | Type: NEGATIVE / Material: negative stain |
Vitrification | Cryogen name: ETHANE |
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Electron microscopy
Microscope | FEI TITAN KRIOS |
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Image recording | Film or detector model: FEI FALCON II (4k x 4k) / Detector mode: INTEGRATING / Average exposure time: 3.0 sec. / Average electron dose: 20.0 e/Å2 Details: Images were stored containing seven parts, where each part represented a set of frames corresponding to a dose of ~20 electrons per Angstrom^2. The full dose image stack was used for the ...Details: Images were stored containing seven parts, where each part represented a set of frames corresponding to a dose of ~20 electrons per Angstrom^2. The full dose image stack was used for the estimation of the CTF as well as for boxing filaments. Only the first two parts were used for the reconstruction (~5 electrons per Angstrom^2). |
Electron beam | Acceleration voltage: 300 kV / Electron source: ![]() |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD |
Experimental equipment | ![]() Model: Titan Krios / Image courtesy: FEI Company |
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Image processing
Final reconstruction | Applied symmetry - Helical parameters - Δz: 27.25 Å Applied symmetry - Helical parameters - Δ&Phi: -166.87 ° Applied symmetry - Helical parameters - Axial symmetry: C1 (asymmetric) Algorithm: BACK PROJECTION / Resolution.type: BY AUTHOR / Resolution: 7.0 Å / Resolution method: OTHER / Software - Name: SPIDER / Details: model-map FSC 0.38 cut-off / Number images used: 12443 |
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Startup model | Type of model: OTHER / Details: low resolution pure actin filament map |
Final angle assignment | Type: NOT APPLICABLE / Software - Name: SPIDER |
-Atomic model buiding 1
Refinement | Space: REAL |
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Output model | ![]() PDB-6anu: |