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- PDB-6anu: Cryo-EM structure of F-actin complexed with the beta-III-spectrin... -
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Basic information
Entry | Database: PDB / ID: 6anu | ||||||||||||
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Title | Cryo-EM structure of F-actin complexed with the beta-III-spectrin actin-binding domain | ||||||||||||
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![]() | STRUCTURAL PROTEIN / actin binding protein / filament | ||||||||||||
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 / 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 / Regulation of MITF-M-dependent genes involved in pigmentation / 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 / cortical actin cytoskeleton / regulation of cyclin-dependent protein serine/threonine kinase activity / adult behavior / maintenance of blood-brain barrier / cortical cytoskeleton / 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 / cell projection / Translocation of SLC2A4 (GLUT4) to the plasma membrane / actin filament / cell motility / 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 / multicellular organism growth / Schaffer collateral - CA1 synapse / 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 | ELECTRON MICROSCOPY / helical reconstruction / negative staining / cryo EM / Resolution: 7 Å | ||||||||||||
![]() | Wang, F. / Orlova, A. / Avery, A.W. / Hays, T.S. / Egelman, E.H. | ||||||||||||
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
Movie |
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Structure viewer | Molecule: ![]() ![]() |
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PDBx/mmCIF format | ![]() | 527.7 KB | Display | ![]() |
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PDB format | ![]() | 417.9 KB | Display | ![]() |
PDBx/mmJSON format | ![]() | Tree view | ![]() | |
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-Validation report
Arichive directory | ![]() ![]() | HTTPS FTP |
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-Related structure data
Related structure data | ![]() 8886MC M: map data used to model this data C: citing same article ( |
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Similar structure data |
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Links
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Assembly
Deposited unit | ![]()
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1 |
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Symmetry | Helical symmetry: (Circular symmetry: 1 / Dyad axis: no / N subunits divisor: 1 / Num. of operations: 20 / Rise per n subunits: 27.25 Å / Rotation per n subunits: -166.87 °) |
Details | THE ASSEMBLY REPRESENTED IN THIS ENTRY HAS REGULAR HELICAL SYMMETRY WITH THE FOLLOWING PARAMETERS: ROTATION PER SUBUNIT (TWIST) = -166.87 DEGREES RISE PER SUBUNIT (HEIGHT) = 27.25 ANGSTROMS |
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Components
#1: Protein | Mass: 41782.660 Da / Num. of mol.: 6 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() ![]() #2: Protein | Mass: 32857.141 Da / Num. of mol.: 6 / Mutation: L253P Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() ![]() |
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-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: FILAMENT / 3D reconstruction method: helical reconstruction |
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Sample preparation
Component | Name: F-actin complexed with the spectrin actin-binding domain Type: COMPLEX / Entity ID: all / Source: RECOMBINANT |
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Source (natural) | Organism: ![]() |
Source (recombinant) | Organism: ![]() ![]() |
Buffer solution | pH: 7.4 |
Specimen | Embedding applied: NO / Shadowing applied: NO / Staining applied: YES / Vitrification applied: YES |
EM staining | Type: NEGATIVE / Material: negative stain |
Vitrification | Cryogen name: ETHANE |
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Electron microscopy imaging
Experimental equipment | ![]() Model: Titan Krios / Image courtesy: FEI Company |
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Microscopy | Model: FEI TITAN KRIOS |
Electron gun | Electron source: ![]() |
Electron lens | Mode: BRIGHT FIELD |
Image recording | Average exposure time: 3 sec. / Electron dose: 20 e/Å2 / Detector mode: INTEGRATING / Film or detector model: FEI FALCON II (4k x 4k) 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). |
Image scans | Movie frames/image: 7 |
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Processing
Software | Name: PHENIX / Version: dev_2471: / Classification: refinement | ||||||||||||||||||||||||||||||
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EM software |
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||||||||
Helical symmerty | Angular rotation/subunit: -166.87 ° / Axial rise/subunit: 27.25 Å / Axial symmetry: C1 | ||||||||||||||||||||||||||||||
3D reconstruction | Resolution: 7 Å / Resolution method: OTHER / Num. of particles: 12443 / Algorithm: BACK PROJECTION / Details: model-map FSC 0.38 cut-off / Symmetry type: HELICAL | ||||||||||||||||||||||||||||||
Atomic model building | Space: REAL | ||||||||||||||||||||||||||||||
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