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- EMDB-21535: CaMKII alpha-30 Cryo-EM reconstruction -

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Basic information

Entry
Database: EMDB / ID: EMD-21535
TitleCaMKII alpha-30 Cryo-EM reconstruction
Map dataMap
Sample
  • Complex: CaMKII alpha-30
    • Protein or peptide: Calcium/calmodulin-dependent protein kinase type II subunit alpha
    • Protein or peptide: Calcium/calmodulin-dependent protein kinase type II subunit alpha
KeywordsCalcium calmodulin dependent protein kinase II / holoenzyme / splicing / SIGNALING PROTEIN / TRANSFERASE
Function / homology
Function and homology information


peptidyl-threonine autophosphorylation / regulation of endocannabinoid signaling pathway / calcium- and calmodulin-dependent protein kinase complex / Ca2+/calmodulin-dependent protein kinase / regulation of neurotransmitter secretion / regulation of neuron migration / dendritic spine development / Trafficking of AMPA receptors / positive regulation of calcium ion transport / negative regulation of hydrolase activity ...peptidyl-threonine autophosphorylation / regulation of endocannabinoid signaling pathway / calcium- and calmodulin-dependent protein kinase complex / Ca2+/calmodulin-dependent protein kinase / regulation of neurotransmitter secretion / regulation of neuron migration / dendritic spine development / Trafficking of AMPA receptors / positive regulation of calcium ion transport / negative regulation of hydrolase activity / Assembly and cell surface presentation of NMDA receptors / calcium/calmodulin-dependent protein kinase activity / regulation of mitochondrial membrane permeability involved in apoptotic process / CaMK IV-mediated phosphorylation of CREB / positive regulation of cardiac muscle cell apoptotic process / Negative regulation of NMDA receptor-mediated neuronal transmission / Phase 0 - rapid depolarisation / Unblocking of NMDA receptors, glutamate binding and activation / regulation of neuronal synaptic plasticity / Ion transport by P-type ATPases / Long-term potentiation / Regulation of MECP2 expression and activity / HSF1-dependent transactivation / glutamate receptor binding / cellular response to interferon-beta / Ion homeostasis / Ras activation upon Ca2+ influx through NMDA receptor / response to ischemia / angiotensin-activated signaling pathway / positive regulation of receptor signaling pathway via JAK-STAT / RAF activation / cellular response to type II interferon / G1/S transition of mitotic cell cycle / Signaling by RAF1 mutants / Signaling by moderate kinase activity BRAF mutants / Paradoxical activation of RAF signaling by kinase inactive BRAF / Signaling downstream of RAS mutants / calcium ion transport / Interferon gamma signaling / endocytic vesicle membrane / Signaling by BRAF and RAF1 fusions / kinase activity / positive regulation of NF-kappaB transcription factor activity / Ca2+ pathway / RAF/MAP kinase cascade / peptidyl-serine phosphorylation / protein autophosphorylation / dendritic spine / postsynaptic density / calmodulin binding / neuron projection / protein phosphorylation / protein serine kinase activity / protein serine/threonine kinase activity / protein homodimerization activity / mitochondrion / nucleoplasm / ATP binding / identical protein binding / nucleus / metal ion 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 / 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. ...Calcium/calmodulin-dependent protein kinase II, association-domain / Calcium/calmodulin dependent protein kinase II association domain / NTF2-like domain superfamily / 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
Similarity search - Component
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 4.8 Å
AuthorsChao LH / Stratton MM
Funding support United States, 1 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R01GM123157 United States
CitationJournal: Sci Signal / Year: 2020
Title: Heterogeneity in human hippocampal CaMKII transcripts reveals allosteric hub-dependent regulation.
Authors: Roman Sloutsky / Noelle Dziedzic / Matthew J Dunn / Rachel M Bates / Ana P Torres-Ocampo / Sivakumar Boopathy / Brendan Page / John G Weeks / Luke H Chao / Margaret M Stratton /
Abstract: Calcium/calmodulin-dependent protein kinase II (CaMKII) plays a central role in Ca signaling throughout the body. In the hippocampus, CaMKII is required for learning and memory. Vertebrate genomes ...Calcium/calmodulin-dependent protein kinase II (CaMKII) plays a central role in Ca signaling throughout the body. In the hippocampus, CaMKII is required for learning and memory. Vertebrate genomes encode four CaMKII homologs: CaMKIIα, CaMKIIβ, CaMKIIγ, and CaMKIIδ. All CaMKIIs consist of a kinase domain, a regulatory segment, a variable linker region, and a hub domain, which is responsible for oligomerization. The four proteins differ primarily in linker length and composition because of extensive alternative splicing. Here, we report the heterogeneity of CaMKII transcripts in three complex samples of human hippocampus using deep sequencing. We showed that hippocampal cells contain a diverse collection of over 70 CaMKII transcripts from all four CaMKII-encoding genes. We characterized the Ca/CaM sensitivity of hippocampal CaMKII variants spanning a broad range of linker lengths and compositions. The effect of the variable linker on Ca/CaM sensitivity depended on the kinase and hub domains. Moreover, we revealed a previously uncharacterized role for the hub domain as an allosteric regulator of kinase activity, which may provide a pharmacological target for modulating CaMKII activity. Using small-angle x-ray scattering and single-particle cryo-electron microscopy (cryo-EM), we present evidence for extensive interactions between the kinase and the hub domains, even in the presence of a 30-residue linker. Together, these data suggest that Ca/CaM sensitivity in CaMKII is homolog dependent and includes substantial contributions from the hub domain. Our sequencing approach, combined with biochemistry, provides insights into understanding the complex pool of endogenous CaMKII splice variants.
History
DepositionMar 11, 2020-
Header (metadata) releaseJul 15, 2020-
Map releaseJul 15, 2020-
UpdateMar 6, 2024-
Current statusMar 6, 2024Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.0072
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by cylindrical radius
  • Surface level: 0.0072
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-6w4o
  • Surface level: 0.0072
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_21535.png

Downloads & links

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Map

FileDownload / File: emd_21535.map.gz / Format: CCP4 / Size: 244.1 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationMap
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.91 Å/pix.
x 400 pix.
= 364. Å		0.91 Å/pix.
x 400 pix.
= 364. Å		0.91 Å/pix.
x 400 pix.
= 364. Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.91 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.0072 / Movie #1: 0.0072
Minimum - Maximum-0.010597348 - 0.026060304
Average (Standard dev.)-0.000005896208 (±0.0010344821)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions400400400
Spacing400400400
CellA=B=C: 364.0 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z0.910.910.91
M x/y/z400400400
origin x/y/z0.0000.0000.000
length x/y/z364.000364.000364.000
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS400400400
D min/max/mean-0.0110.026-0.000

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Supplemental data

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Mask #1

Fileemd_21535_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: half-map 1

Fileemd_21535_half_map_1.map
Annotationhalf-map 1
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: half-map 2

Fileemd_21535_half_map_2.map
Annotationhalf-map 2
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : CaMKII alpha-30

EntireName: CaMKII alpha-30
Components
  • Complex: CaMKII alpha-30
    • Protein or peptide: Calcium/calmodulin-dependent protein kinase type II subunit alpha
    • Protein or peptide: Calcium/calmodulin-dependent protein kinase type II subunit alpha

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Supramolecule #1: CaMKII alpha-30

SupramoleculeName: CaMKII alpha-30 / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Homo sapiens (human)

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Macromolecule #1: Calcium/calmodulin-dependent protein kinase type II subunit alpha

MacromoleculeName: Calcium/calmodulin-dependent protein kinase type II subunit alpha
type: protein_or_peptide / ID: 1 / Number of copies: 12 / Enantiomer: LEVO / EC number: Ca2+/calmodulin-dependent protein kinase
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 53.633953 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: STRFTEEYQL FEELGKGAFS VVRRCVKVLA GQEYAAKIIN TKKLSARDHQ KLEREARICR LLKHPNIVRL HDSISEEGHH YLIFDLVTG GELFEDIVAR EYYSEADASH CIQQILEAVL HCHQMGVVHR DLKPENLLLA SKLKGAAVKL ADFGLAIEVE G EQQAWFGF ...String:
STRFTEEYQL FEELGKGAFS VVRRCVKVLA GQEYAAKIIN TKKLSARDHQ KLEREARICR LLKHPNIVRL HDSISEEGHH YLIFDLVTG GELFEDIVAR EYYSEADASH CIQQILEAVL HCHQMGVVHR DLKPENLLLA SKLKGAAVKL ADFGLAIEVE G EQQAWFGF AGTPGYLSPE VLRKDPYGKP VDLWACGVIL YILLVGYPPF WDEDQHRLYQ QIKAGAYDFP SPEWDTVTPE AK DLINKML TINPSKRITA AEALKHPWIS HRSTVASCMH RQETVDCLKK FNARRKLKGA ILTTMLATRN FSGGKSGGNK KSD GVKESS ESTNTTIEDE DTKVRKQEII KVTEQLIEAI SNGDFESYTK MCDPGMTAFE PEALGNLVEG LDFHRFYFEN LWSR NSKPV HTTILNPHIH LMGDESACIA YIRITQYLDA GGIPRTAQSE ETRVWHRRDG KWQIVHFHRS GAPSVLPH

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

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Macromolecule #2: Calcium/calmodulin-dependent protein kinase type II subunit alpha

MacromoleculeName: Calcium/calmodulin-dependent protein kinase type II subunit alpha
type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO / EC number: Ca2+/calmodulin-dependent protein kinase
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 53.634984 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: STRFTEEYQL FEELGKGAFS VVRRCVKVLA GQEYAAMIIN TKKLSARDHQ KLEREARICR LLKHPNIVRL HDSISEEGHH YLIFDLVTG GELFEDIVAR EYYSEADASH CIQQILEAVL HCHQMGVVHR NLKPENLLLA SKLKGAAVKL ADFGLAIEVE G EQQAWFGF ...String:
STRFTEEYQL FEELGKGAFS VVRRCVKVLA GQEYAAMIIN TKKLSARDHQ KLEREARICR LLKHPNIVRL HDSISEEGHH YLIFDLVTG GELFEDIVAR EYYSEADASH CIQQILEAVL HCHQMGVVHR NLKPENLLLA SKLKGAAVKL ADFGLAIEVE G EQQAWFGF AGTPGYLSPE VLRKDPYGKP VDLWACGVIL YILLVGYPPF WDEDQHRLYQ QIKAGAYDFP SPEWDTVTPE AK DLINKML TINPSKRITA AEALKHPWIS HRSTVASCMH RQETVDCLKK FNARRKLKGA ILTTMLATRN FSGGKSGGNK KSD GVKESS ESTNTTIEDE DTKVRKQEII KVTEQLIEAI SNGDFESYTK MCDPGMTAFE PEALGNLVEG LDFHRFYFEN LWSR NSKPV HTTILNPHIH LMGDESACIA YIRITQYLDA GGIPRTAQSE ETRVWHRRDG KWQIVHFHRS GAPSVLPH

UniProtKB: 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

BufferpH: 8
GridModel: C-flat-1.2/1.3 / Material: GOLD / Mesh: 300 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 30 sec. / Details: 30 mA for 30 s using PELCO easiGlow
VitrificationCryogen name: ETHANE / Chamber humidity: 96 % / Instrument: GATAN CRYOPLUNGE 3

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Electron microscopy

MicroscopeFEI TALOS ARCTICA
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: SUPER-RESOLUTION / Average electron dose: 52.0 e/Å2
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: OTHER
Experimental equipment
Model: Talos Arctica / Image courtesy: FEI Company

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Image processing

Startup modelType of model: OTHER / Details: RELION Initial Model
Final reconstructionResolution.type: BY AUTHOR / Resolution: 4.8 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION / Number images used: 160211
Initial angle assignmentType: RANDOM ASSIGNMENT / Software - Name: RELION
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION
Final 3D classificationSoftware - Name: RELION
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial model
PDB IDChain

3soa

source_name: PDB, initial_model_type: experimental model

5ig3

source_name: PDB, initial_model_type: experimental model
RefinementProtocol: RIGID BODY FIT
Output model

PDB-6w4o:
CaMKII alpha-30 Cryo-EM reconstruction

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