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- EMDB-5756: 3D EM Reconstruction of the AKAP18-PKA Complex in a Linear Confor... -
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Basic information
Entry | Database: EMDB / ID: EMD-5756 | |||||||||
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Title | 3D EM Reconstruction of the AKAP18-PKA Complex in a Linear Conformation | |||||||||
![]() | Reconstruction of AKAP18-PKA in a Linear Conformation | |||||||||
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![]() | A-Kinase Anchoring Protein / AKAP / AKAP18 / A-Kinase / PKA / cAMP-Dependent Kinase / Kinase / RII / PKA Regulatory Subunit II / Phosphorylation / Anchoring / Intrinsic Disorder | |||||||||
Function / homology | ![]() spontaneous exocytosis of neurotransmitter / PKA activation in glucagon signalling / CREB1 phosphorylation through the activation of Adenylate Cyclase / negative regulation of meiotic cell cycle / HDL assembly / DARPP-32 events / Rap1 signalling / PKA activation / Vasopressin regulates renal water homeostasis via Aquaporins / Regulation of insulin secretion ...spontaneous exocytosis of neurotransmitter / PKA activation in glucagon signalling / CREB1 phosphorylation through the activation of Adenylate Cyclase / negative regulation of meiotic cell cycle / HDL assembly / DARPP-32 events / Rap1 signalling / PKA activation / Vasopressin regulates renal water homeostasis via Aquaporins / Regulation of insulin secretion / GPER1 signaling / Hedgehog 'off' state / Glucagon-like Peptide-1 (GLP1) regulates insulin secretion / exocytic vesicle / Loss of Nlp from mitotic centrosomes / Recruitment of mitotic centrosome proteins and complexes / Loss of proteins required for interphase microtubule organization from the centrosome / regulation of membrane repolarization / Recruitment of NuMA to mitotic centrosomes / Anchoring of the basal body to the plasma membrane / AURKA Activation by TPX2 / MAPK6/MAPK4 signaling / GLI3 is processed to GLI3R by the proteasome / Factors involved in megakaryocyte development and platelet production / cAMP-dependent protein kinase regulator activity / Regulation of PLK1 Activity at G2/M Transition / Interleukin-3, Interleukin-5 and GM-CSF signaling / CD209 (DC-SIGN) signaling / Mitochondrial protein degradation / RET signaling / positive regulation of potassium ion transmembrane transport / nucleotide-activated protein kinase complex / regulation of protein kinase A signaling / Ion homeostasis / VEGFA-VEGFR2 Pathway / regulation of cellular respiration / regulation of protein processing / protein localization to lipid droplet / regulation of bicellular tight junction assembly / cellular response to parathyroid hormone stimulus / cAMP-dependent protein kinase inhibitor activity / beta-2 adrenergic receptor binding / cAMP-dependent protein kinase / cellular response to cold / regulation of osteoblast differentiation / sperm capacitation / protein kinase A binding / cAMP-dependent protein kinase activity / ciliary base / negative regulation of glycolytic process through fructose-6-phosphate / cAMP-dependent protein kinase complex / AMP-activated protein kinase activity / postsynaptic modulation of chemical synaptic transmission / cellular response to glucagon stimulus / AMP binding / protein kinase A regulatory subunit binding / small molecule binding / plasma membrane raft / protein kinase A catalytic subunit binding / axoneme / mesoderm formation / lateral plasma membrane / sperm flagellum / regulation of proteasomal protein catabolic process / negative regulation of smoothened signaling pathway / cAMP binding / cellular response to cAMP / positive regulation of gluconeogenesis / sperm midpiece / negative regulation of TORC1 signaling / regulation of synaptic transmission, glutamatergic / T-tubule / protein kinase A signaling / protein serine/threonine/tyrosine kinase activity / protein export from nucleus / hippocampal mossy fiber to CA3 synapse / phosphorylation / acrosomal vesicle / positive regulation of protein export from nucleus / sarcoplasmic reticulum / neural tube closure / cellular response to glucose stimulus / regulation of protein phosphorylation / modulation of chemical synaptic transmission / neuromuscular junction / positive regulation of insulin secretion / adenylate cyclase-activating G protein-coupled receptor signaling pathway / small GTPase binding / mRNA processing / presynapse / kinase activity / manganese ion binding / cellular response to heat / peptidyl-serine phosphorylation / postsynapse / dendritic spine / regulation of cell cycle / protein kinase activity / nuclear speck / apical plasma membrane Similarity search - Function | |||||||||
Biological species | ![]() ![]() ![]() | |||||||||
Method | single particle reconstruction / negative staining / Resolution: 35.0 Å | |||||||||
![]() | Smith FD / Reichow SL / Esseltine JL / Shi D / Langeberg LK / Scott JD / Gonen T | |||||||||
![]() | ![]() Title: Intrinsic disorder within an AKAP-protein kinase A complex guides local substrate phosphorylation. Authors: F Donelson Smith / Steve L Reichow / Jessica L Esseltine / Dan Shi / Lorene K Langeberg / John D Scott / Tamir Gonen / ![]() Abstract: Anchoring proteins sequester kinases with their substrates to locally disseminate intracellular signals and avert indiscriminate transmission of these responses throughout the cell. Mechanistic ...Anchoring proteins sequester kinases with their substrates to locally disseminate intracellular signals and avert indiscriminate transmission of these responses throughout the cell. Mechanistic understanding of this process is hampered by limited structural information on these macromolecular complexes. A-kinase anchoring proteins (AKAPs) spatially constrain phosphorylation by cAMP-dependent protein kinases (PKA). Electron microscopy and three-dimensional reconstructions of type-II PKA-AKAP18γ complexes reveal hetero-pentameric assemblies that adopt a range of flexible tripartite configurations. Intrinsically disordered regions within each PKA regulatory subunit impart the molecular plasticity that affords an ∼16 nanometer radius of motion to the associated catalytic subunits. Manipulating flexibility within the PKA holoenzyme augmented basal and cAMP responsive phosphorylation of AKAP-associated substrates. Cell-based analyses suggest that the catalytic subunit remains within type-II PKA-AKAP18γ complexes upon cAMP elevation. We propose that the dynamic movement of kinase sub-structures, in concert with the static AKAP-regulatory subunit interface, generates a solid-state signaling microenvironment for substrate phosphorylation. DOI: http://dx.doi.org/10.7554/eLife.01319.001. | |||||||||
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 | ![]() | 6.1 MB | ![]() | |
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Header (meta data) | ![]() ![]() | 15.1 KB 15.1 KB | Display Display | ![]() |
Images | ![]() | 61.5 KB | ||
Archive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 3j4rMC ![]() 5755C ![]() 3j4qC 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 | Reconstruction of AKAP18-PKA in a Linear Conformation | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 4.2 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 : AKAP18-PKA Complex in Linear Conformation
Entire | Name: AKAP18-PKA Complex in Linear Conformation |
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Components |
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-Supramolecule #1000: AKAP18-PKA Complex in Linear Conformation
Supramolecule | Name: AKAP18-PKA Complex in Linear Conformation / type: sample / ID: 1000 Oligomeric state: Hetero-pentamer composed of one AKAP18 bound to a dimer of the PKA Regulatory Subunit II and two copies of the PKA Catalytic Subunit Number unique components: 3 |
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Molecular weight | Experimental: 240 KDa / Theoretical: 220 KDa / Method: Native PAGE |
-Macromolecule #1: A-Kinase Anchoring Protein 18
Macromolecule | Name: A-Kinase Anchoring Protein 18 / type: protein_or_peptide / ID: 1 / Name.synonym: AKAP18 / Number of copies: 1 / Oligomeric state: monomer / Recombinant expression: Yes |
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Source (natural) | Organism: ![]() |
Molecular weight | Theoretical: 40 KDa |
Recombinant expression | Organism: ![]() ![]() |
-Macromolecule #2: A-Protein Kinase Catalytic Subunit
Macromolecule | Name: A-Protein Kinase Catalytic Subunit / type: protein_or_peptide / ID: 2 / Name.synonym: PKA C subunit / Number of copies: 2 / Oligomeric state: monomer / Recombinant expression: Yes |
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Source (natural) | Organism: ![]() ![]() |
Molecular weight | Theoretical: 40 KDa |
Recombinant expression | Organism: ![]() ![]() |
-Macromolecule #3: A-Protein Kinase Regulatory Subunit II alpha
Macromolecule | Name: A-Protein Kinase Regulatory Subunit II alpha / type: protein_or_peptide / ID: 3 / Name.synonym: PKA RIIalpha Subunit / Number of copies: 2 / Oligomeric state: dimer / Recombinant expression: Yes |
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Source (natural) | Organism: ![]() ![]() |
Molecular weight | Theoretical: 45 KDa |
Recombinant expression | Organism: ![]() ![]() |
-Experimental details
-Structure determination
Method | negative staining |
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![]() | single particle reconstruction |
Aggregation state | particle |
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Sample preparation
Concentration | 0.01 mg/mL |
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Buffer | pH: 7.4 Details: 25 mM HEPES, 200 mM NaCl, 0.5 mM EDTA, 1 mM dithiothreitol. |
Staining | Type: NEGATIVE / Details: 0.75% (w/v) uranyl formate |
Grid | Details: 200 mesh copper grid with thin carbon support |
Vitrification | Cryogen name: NONE / Instrument: OTHER |
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Electron microscopy
Microscope | FEI TECNAI 12 |
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Temperature | Average: 298 K |
Alignment procedure | Legacy - Astigmatism: Objective lens astigmatism was corrected at 100,000 times magnification |
Date | May 1, 2012 |
Image recording | Category: CCD / Film or detector model: GENERIC GATAN / Number real images: 1335 / Average electron dose: 15 e/Å2 |
Tilt angle min | 0 |
Electron beam | Acceleration voltage: 120 kV / Electron source: LAB6 |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 6.3 mm / Nominal defocus max: 2.5 µm / Nominal defocus min: 1.0 µm / Nominal magnification: 52000 |
Sample stage | Specimen holder model: OTHER / Tilt angle max: 50 |
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Image processing
Details | Images were processed in IMAGIC and ISAC. 3D reconstruction was done in IMAGIC and FREALIGN. |
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CTF correction | Details: Each Micrograph |
Final reconstruction | Resolution.type: BY AUTHOR / Resolution: 35.0 Å / Resolution method: FSC 0.5 CUT-OFF / Software - Name: IMAGIC, FREALIGN / Number images used: 1000 |
-Atomic model buiding 1
Initial model | PDB ID: Chain - #0 - Chain ID: A / Chain - #1 - Chain ID: B |
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Software | Name: ![]() |
Details | This domain was linked to the AKAP18 central domain (PDB 2VFL) using COOT and they were fit together into the EM density. |
Refinement | Space: REAL / Protocol: RIGID BODY FIT / Target criteria: cross-correlation |
Output model | ![]() PDB-3j4r: |
-Atomic model buiding 2
Initial model | PDB ID: Chain - Chain ID: A |
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Software | Name: ![]() |
Details | This domain was linked to the AKAP RII Binding Domain (PDB 2IZX) using COOT and they were fit together into the EM density. |
Refinement | Space: REAL / Protocol: RIGID BODY FIT / Target criteria: cross-correlation |
Output model | ![]() PDB-3j4r: |
-Atomic model buiding 3
Initial model | PDB ID: Chain - #0 - Chain ID: A / Chain - #1 - Chain ID: B |
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Software | Name: ![]() |
Details | This domain was linked to the PKA RIIalpha D/D domain (PDB 2IZX) using COOT and they were fit together into the EM density. |
Refinement | Space: REAL / Protocol: RIGID BODY FIT / Target criteria: cross-correlation |
Output model | ![]() PDB-3j4r: |