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- EMDB-40955: Cryo-EM structure of dodecameric hub domain of CaMKII alpha -

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Basic information

Entry
Database: EMDB / ID: EMD-40955
TitleCryo-EM structure of dodecameric hub domain of CaMKII alpha
Map dataDodecameric CaMKII alpha hub
Sample
  • Complex: Venus-tagged CaMKII Alpha Association Domain
    • Protein or peptide: Venus-tagged CaMKII Alpha Association Domain
KeywordsHigh-order oligomer / Protein Kinase / Signaling / Memory / SIGNALING PROTEIN
Function / homology
Function and homology information


regulation of synaptic vesicle docking / HSF1-dependent transactivation / RAF activation / Ion transport by P-type ATPases / peptidyl-threonine autophosphorylation / regulation of endocannabinoid signaling pathway / calcium- and calmodulin-dependent protein kinase complex / neurotransmitter receptor transport to plasma membrane / Interferon gamma signaling / Ca2+/calmodulin-dependent protein kinase ...regulation of synaptic vesicle docking / HSF1-dependent transactivation / RAF activation / Ion transport by P-type ATPases / peptidyl-threonine autophosphorylation / regulation of endocannabinoid signaling pathway / calcium- and calmodulin-dependent protein kinase complex / neurotransmitter receptor transport to plasma membrane / Interferon gamma signaling / Ca2+/calmodulin-dependent protein kinase / negative regulation of hydrolase activity / dendritic spine development / regulation of neurotransmitter secretion / Trafficking of AMPA receptors / positive regulation of calcium ion transport / regulation of neuron migration / GTPase activating protein binding / Ca2+ pathway / calcium/calmodulin-dependent protein kinase activity / RAF/MAP kinase cascade / regulation of mitochondrial membrane permeability involved in apoptotic process / Ion homeostasis / dendrite morphogenesis / regulation of neurotransmitter receptor localization to postsynaptic specialization membrane / regulation of neuronal synaptic plasticity / glutamate receptor binding / Unblocking of NMDA receptors, glutamate binding and activation / postsynaptic cytosol / regulation of protein localization to plasma membrane / cellular response to interferon-beta / positive regulation of cardiac muscle cell apoptotic process / presynaptic cytosol / ionotropic glutamate receptor signaling pathway / dendrite cytoplasm / bioluminescence / response to ischemia / angiotensin-activated signaling pathway / generation of precursor metabolites and energy / positive regulation of receptor signaling pathway via JAK-STAT / G1/S transition of mitotic cell cycle / cellular response to type II interferon / Schaffer collateral - CA1 synapse / calcium ion transport / kinase activity / dendritic spine / calmodulin binding / neuron projection / postsynaptic density / axon / protein serine kinase activity / neuronal cell body / protein serine/threonine kinase activity / synapse / dendrite / glutamatergic synapse / protein homodimerization activity / mitochondrion / ATP binding / metal ion binding / identical protein binding / cytosol / cytoplasm
Similarity search - Function
Calcium/calmodulin-dependent protein kinase II, association-domain / Calcium/calmodulin dependent protein kinase II association domain / NTF2-like domain superfamily / Green fluorescent protein, GFP / Green fluorescent protein-related / Green fluorescent protein / Green fluorescent protein / Serine/threonine-protein kinase, active site / Serine/Threonine protein kinases active-site signature. / Protein kinase domain ...Calcium/calmodulin-dependent protein kinase II, association-domain / Calcium/calmodulin dependent protein kinase II association domain / NTF2-like domain superfamily / Green fluorescent protein, GFP / Green fluorescent protein-related / Green fluorescent protein / Green fluorescent protein / Serine/threonine-protein kinase, active site / Serine/Threonine protein kinases active-site signature. / Protein kinase domain / Serine/Threonine protein kinases, catalytic domain / Protein kinase, ATP binding site / Protein kinases ATP-binding region signature. / Protein kinase domain profile. / Protein kinase domain / Protein kinase-like domain superfamily
Similarity search - Domain/homology
Calcium/calmodulin-dependent protein kinase type II subunit alpha / Green fluorescent protein
Similarity search - Component
Biological speciesRattus norvegicus (Norway rat)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.7 Å
AuthorsChien C-T / Chiu W / Khan S
Funding support United States, 1 items
OrganizationGrant numberCountry
Other governmentU24 GM129541 United States
CitationJournal: Commun Biol / Year: 2024
Title: Hub stability in the calcium calmodulin-dependent protein kinase II.
Authors: Chih-Ta Chien / Henry Puhl / Steven S Vogel / Justin E Molloy / Wah Chiu / Shahid Khan /
Abstract: The calcium calmodulin protein kinase II (CaMKII) is a multi-subunit ring assembly with a central hub formed by the association domains. There is evidence for hub polymorphism between and within ...The calcium calmodulin protein kinase II (CaMKII) is a multi-subunit ring assembly with a central hub formed by the association domains. There is evidence for hub polymorphism between and within CaMKII isoforms, but the link between polymorphism and subunit exchange has not been resolved. Here, we present near-atomic resolution cryogenic electron microscopy (cryo-EM) structures revealing that hubs from the α and β isoforms, either standalone or within an β holoenzyme, coexist as 12 and 14 subunit assemblies. Single-molecule fluorescence microscopy of Venus-tagged holoenzymes detects intermediate assemblies and progressive dimer loss due to intrinsic holoenzyme lability, and holoenzyme disassembly into dimers upon mutagenesis of a conserved inter-domain contact. Molecular dynamics (MD) simulations show the flexibility of 4-subunit precursors, extracted in-silico from the β hub polymorphs, encompassing the curvature of both polymorphs. The MD explains how an open hub structure also obtained from the β holoenzyme sample could be created by dimer loss and analysis of its cryo-EM dataset reveals how the gap could open further. An assembly model, considering dimer concentration dependence and strain differences between polymorphs, proposes a mechanism for intrinsic hub lability to fine-tune the stoichiometry of αβ heterooligomers for their dynamic localization within synapses in neurons.
History
DepositionJun 1, 2023-
Header (metadata) releaseJun 12, 2024-
Map releaseJun 12, 2024-
UpdateJul 10, 2024-
Current statusJul 10, 2024Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_40955.png

Downloads & links

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Map

FileDownload / File: emd_40955.map.gz / Format: CCP4 / Size: 216 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationDodecameric CaMKII alpha hub
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.74 Å/pix.
x 384 pix.
= 284.544 Å		0.74 Å/pix.
x 384 pix.
= 284.544 Å		0.74 Å/pix.
x 384 pix.
= 284.544 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.741 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.5
Minimum - Maximum-3.0573993 - 5.5736704
Average (Standard dev.)0.0015362559 (±0.096780695)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions384384384
Spacing384384384
CellA=B=C: 284.544 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #2

Fileemd_40955_half_map_1.map
Projections & Slices
AxesZYX

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Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #1

Fileemd_40955_half_map_2.map
Projections & Slices
AxesZYX

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Histogram (log scale)

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Sample components

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Entire : Venus-tagged CaMKII Alpha Association Domain

EntireName: Venus-tagged CaMKII Alpha Association Domain
Components
  • Complex: Venus-tagged CaMKII Alpha Association Domain
    • Protein or peptide: Venus-tagged CaMKII Alpha Association Domain

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Supramolecule #1: Venus-tagged CaMKII Alpha Association Domain

SupramoleculeName: Venus-tagged CaMKII Alpha Association Domain / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Rattus norvegicus (Norway rat)

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Macromolecule #1: Venus-tagged CaMKII Alpha Association Domain

MacromoleculeName: Venus-tagged CaMKII Alpha Association Domain / type: protein_or_peptide / ID: 1 / Number of copies: 12 / Enantiomer: LEVO / EC number: Ca2+/calmodulin-dependent protein kinase
Source (natural)Organism: Rattus norvegicus (Norway rat)
Molecular weightTheoretical: 45.877539 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MGSSHHHHHH SSGLVPRGSH VSKGEELFTG VVPILVELDG DVNGHKFSVS GEGEGDATYG KLTLKLICTT GKLPVPWPTL VTTLGYGLQ CFARYPDHMK QHDFFKSAMP EGYVQERTIF FKDDGNYKTR AEVKFEGDTL VNRIELKGID FKEDGNILGH K LEYNYNSH ...String:
MGSSHHHHHH SSGLVPRGSH VSKGEELFTG VVPILVELDG DVNGHKFSVS GEGEGDATYG KLTLKLICTT GKLPVPWPTL VTTLGYGLQ CFARYPDHMK QHDFFKSAMP EGYVQERTIF FKDDGNYKTR AEVKFEGDTL VNRIELKGID FKEDGNILGH K LEYNYNSH NVYITADKQK NGIKANFKIR HNIEDGGVQL ADHYQQNTPI GDGPVLLPDN HYLSYQSKLS KDPNEKRDHM VL LEFVTAA GITLGMDELY KSGLRSRAQA SNSAVDVRKQ EIIKVTEQLI EAISNGDFES YTKMCDPGMT AFEPEALGNL VEG LDFHRF YFENLWSRNS KPVHTTILNP HIHLMGDESA CIAYIRITQY LDAGGIPRTA QSEETRVWHR RDGKWQIVHF HRSG APSVL PH

UniProtKB: Green fluorescent protein, Calcium/calmodulin-dependent protein kinase type II subunit alpha

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

Concentration20 mg/mL
BufferpH: 7.5
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: FEI FALCON IV (4k x 4k) / Average electron dose: 60.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 1.5 µm / Nominal defocus min: 0.6 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: INSILICO MODEL
Final reconstructionResolution.type: BY AUTHOR / Resolution: 2.7 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 94506
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD

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