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- EMDB-47944: Reset Type-I Protein Kinase A Holoenzyme -

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Basic information

Entry
Database: EMDB / ID: EMD-47944
TitleReset Type-I Protein Kinase A Holoenzyme
Map data
Sample
  • Complex: Reset type-I PKA holoenzyme complex of regulatory and catalytic subunits
    • Protein or peptide: cAMP-dependent protein kinase catalytic subunit alpha
    • Protein or peptide: cAMP-dependent protein kinase type I-alpha regulatory subunit
  • Ligand: ADENOSINE-5'-TRIPHOSPHATE
KeywordsKinase / Regulator / SIGNALING PROTEIN
Function / homology
Function and homology information


PKA-mediated phosphorylation of CREB / PKA-mediated phosphorylation of key metabolic factors / sperm head-tail coupling apparatus / ROBO receptors bind AKAP5 / channel activator activity / HDL assembly / PKA activation in glucagon signalling / DARPP-32 events / CREB1 phosphorylation through the activation of Adenylate Cyclase / GPER1 signaling ...PKA-mediated phosphorylation of CREB / PKA-mediated phosphorylation of key metabolic factors / sperm head-tail coupling apparatus / ROBO receptors bind AKAP5 / channel activator activity / HDL assembly / PKA activation in glucagon signalling / DARPP-32 events / CREB1 phosphorylation through the activation of Adenylate Cyclase / GPER1 signaling / Factors involved in megakaryocyte development and platelet production / Regulation of glycolysis by fructose 2,6-bisphosphate metabolism / mitochondrial protein catabolic process / PKA activation / nucleotide-activated protein kinase complex / Hedgehog 'off' state / cell communication by electrical coupling involved in cardiac conduction / high-density lipoprotein particle assembly / Rap1 signalling / negative regulation of cAMP/PKA signal transduction / cAMP-dependent protein kinase inhibitor activity / cAMP-dependent protein kinase / regulation of protein processing / Loss of phosphorylation of MECP2 at T308 / cardiac muscle cell proliferation / cAMP-dependent protein kinase activity / CREB1 phosphorylation through the activation of Adenylate Cyclase / PKA activation / protein localization to lipid droplet / cAMP-dependent protein kinase complex / regulation of bicellular tight junction assembly / cellular response to parathyroid hormone stimulus / sarcomere organization / negative regulation of interleukin-2 production / cellular response to cold / regulation of osteoblast differentiation / sperm capacitation / High laminar flow shear stress activates signaling by PIEZO1 and PECAM1:CDH5:KDR in endothelial cells / Vasopressin regulates renal water homeostasis via Aquaporins / negative regulation of glycolytic process through fructose-6-phosphate / ciliary base / Triglyceride catabolism / protein kinase A regulatory subunit binding / negative regulation of activated T cell proliferation / protein kinase A catalytic subunit binding / cAMP/PKA signal transduction / mesoderm formation / RET signaling / PKA activation in glucagon signalling / Regulation of MECP2 expression and activity / Interleukin-3, Interleukin-5 and GM-CSF signaling / sperm flagellum / immunological synapse / DARPP-32 events / plasma membrane raft / axoneme / regulation of cardiac conduction / regulation of cardiac muscle contraction / regulation of macroautophagy / postsynaptic modulation of chemical synaptic transmission / regulation of cardiac muscle contraction by regulation of the release of sequestered calcium ion / renal water homeostasis / Hedgehog 'off' state / cAMP binding / Ion homeostasis / negative regulation of TORC1 signaling / regulation of proteasomal protein catabolic process / cellular response to epinephrine stimulus / sperm midpiece / negative regulation of smoothened signaling pathway / calcium channel complex / Mitochondrial protein degradation / positive regulation of gluconeogenesis / multivesicular body / Loss of Nlp from mitotic centrosomes / Loss of proteins required for interphase microtubule organization from the centrosome / Recruitment of mitotic centrosome proteins and complexes / CD209 (DC-SIGN) signaling / protein serine/threonine/tyrosine kinase activity / cellular response to glucagon stimulus / Recruitment of NuMA to mitotic centrosomes / positive regulation of calcium-mediated signaling / FCGR3A-mediated IL10 synthesis / Anchoring of the basal body to the plasma membrane / regulation of heart rate / protein export from nucleus / acrosomal vesicle / AURKA Activation by TPX2 / positive regulation of protein export from nucleus / Regulation of insulin secretion / neural tube closure / Degradation of GLI1 by the proteasome / positive regulation of cholesterol biosynthetic process / Degradation of GLI2 by the proteasome / GLI3 is processed to GLI3R by the proteasome / cellular response to glucose stimulus / MAPK6/MAPK4 signaling / adenylate cyclase-inhibiting G protein-coupled receptor signaling pathway / neuromuscular junction / positive regulation of insulin secretion
Similarity search - Function
cAMP-dependent protein kinase regulatory subunit / cAMP-dependent protein kinase regulatory subunit, dimerization-anchoring domain / Regulatory subunit of type II PKA R-subunit / RIIalpha, Regulatory subunit portion of type II PKA R-subunit / : / Cyclic nucleotide-binding domain signature 2. / Cyclic nucleotide-binding domain signature 1. / cAMP-dependent protein kinase catalytic subunit / Cyclic nucleotide-binding, conserved site / Cyclic nucleotide-monophosphate binding domain ...cAMP-dependent protein kinase regulatory subunit / cAMP-dependent protein kinase regulatory subunit, dimerization-anchoring domain / Regulatory subunit of type II PKA R-subunit / RIIalpha, Regulatory subunit portion of type II PKA R-subunit / : / Cyclic nucleotide-binding domain signature 2. / Cyclic nucleotide-binding domain signature 1. / cAMP-dependent protein kinase catalytic subunit / Cyclic nucleotide-binding, conserved site / Cyclic nucleotide-monophosphate binding domain / Cyclic nucleotide-binding domain / cAMP/cGMP binding motif profile. / Cyclic nucleotide-binding domain / Cyclic nucleotide-binding domain superfamily / Extension to Ser/Thr-type protein kinases / AGC-kinase, C-terminal / AGC-kinase C-terminal domain profile. / RmlC-like jelly roll fold / 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
cAMP-dependent protein kinase type I-alpha regulatory subunit / cAMP-dependent protein kinase catalytic subunit alpha
Similarity search - Component
Biological speciesHomo sapiens (human) / Bos taurus (domestic cattle)
Methodsingle particle reconstruction / cryo EM / Resolution: 4.18 Å
AuthorsVenkatakrishnan V / Buckley T / Laremore TN / Armache JP / Anand GS
Funding support United States, 1 items
OrganizationGrant numberCountry
Not funded United States
CitationJournal: J Am Chem Soc / Year: 2025
Title: Multiplicity of Regulatory Subunit Conformations Defines Structural Ensemble of Reset Protein Kinase A Holoenzyme.
Authors: Varun Venkatakrishnan / Tatiana N Laremore / Theresa S C Buckley / Jean-Paul Armache / Ganesh S Anand /
Abstract: How protein kinase A (PKA) is reset to a basal state following 3'5'-cyclic adenosine monophosphate (cAMP)-mediated activation is unknown. Here we describe the mechanism of cAMP-PKA type I signal ...How protein kinase A (PKA) is reset to a basal state following 3'5'-cyclic adenosine monophosphate (cAMP)-mediated activation is unknown. Here we describe the mechanism of cAMP-PKA type I signal termination leading to a reset of PKA by holoenzyme formation through the obligatory action of phosphodiesterases (PDEs). We report a catalytic subunit (Cα)-assisted mechanism for the reset of type I PKA and describe for the first time multiple structures of the reset PKA holoenzyme (RIα:Cα) that capture an ensemble of multiple conformational end-states through integrative electron microscopy and structural mass spectrometry approaches. Together these complementary methods highlight the large conformational dynamics of the regulatory subunit (RIα) within the tetrameric reset PKA holoenzyme. The cAMP-free reset PKA holoenzyme adopts multiple distinct conformations of RIα with contributions from the N-terminal linker and CNB-B dynamics. Our findings highlight the interplay between RIα, Cα, and PDEs (PDE8) in cAMP-PKA signalosomes to offer a new paradigm for PDE-mediated regulation of cAMP-PKA signaling.
History
DepositionNov 16, 2024-
Header (metadata) releaseApr 23, 2025-
Map releaseApr 23, 2025-
UpdateApr 23, 2025-
Current statusApr 23, 2025Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_47944.png

Downloads & links

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Map

FileDownload / File: emd_47944.map.gz / Format: CCP4 / Size: 18.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.94 Å/pix.
x 170 pix.
= 160.48 Å		0.94 Å/pix.
x 170 pix.
= 160.48 Å		0.94 Å/pix.
x 170 pix.
= 160.48 Å

Surface

Projections

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Images are generated by Spider.

Voxel sizeX=Y=Z: 0.944 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.13
Minimum - Maximum-1.020253 - 2.113989
Average (Standard dev.)0.00017311689 (±0.049453147)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions170170170
Spacing170170170
CellA=B=C: 160.48 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #2

Fileemd_47944_half_map_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #1

Fileemd_47944_half_map_2.map
Projections & Slices
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Histogram (log scale)

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Sample components

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Entire : Reset type-I PKA holoenzyme complex of regulatory and catalytic s...

EntireName: Reset type-I PKA holoenzyme complex of regulatory and catalytic subunits
Components
  • Complex: Reset type-I PKA holoenzyme complex of regulatory and catalytic subunits
    • Protein or peptide: cAMP-dependent protein kinase catalytic subunit alpha
    • Protein or peptide: cAMP-dependent protein kinase type I-alpha regulatory subunit
  • Ligand: ADENOSINE-5'-TRIPHOSPHATE

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Supramolecule #1: Reset type-I PKA holoenzyme complex of regulatory and catalytic s...

SupramoleculeName: Reset type-I PKA holoenzyme complex of regulatory and catalytic subunits
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#2
Source (natural)Organism: Homo sapiens (human)

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Macromolecule #1: cAMP-dependent protein kinase catalytic subunit alpha

MacromoleculeName: cAMP-dependent protein kinase catalytic subunit alpha / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO / EC number: cAMP-dependent protein kinase
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 40.737297 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: GNAAAAKKGS EQESVKEFLA KAKEDFLKKW ETPSQNTAQL DQFDRIKTLG TGSFGRVMLV KHKESGNHYA MKILDKQKVV KLKQIEHTL NEKRILQAVN FPFLVKLEFS FKDNSNLYMV MEYVAGGEMF SHLRRIGRF(SEP) EPHARFYAAQ IVLTFEY LH SLDLIYRDLK ...String:
GNAAAAKKGS EQESVKEFLA KAKEDFLKKW ETPSQNTAQL DQFDRIKTLG TGSFGRVMLV KHKESGNHYA MKILDKQKVV KLKQIEHTL NEKRILQAVN FPFLVKLEFS FKDNSNLYMV MEYVAGGEMF SHLRRIGRF(SEP) EPHARFYAAQ IVLTFEY LH SLDLIYRDLK PENLLIDQQG YIQVTDFGFA KRVKGRTW(TPO)L CGTPEYLAPE IILSKGYNKA VDWWALGVLI YEMA AGYPP FFADQPIQIY EKIVSGKVRF PSHFSSDLKD LLRNLLQVDL TKRFGNLKNG VNDIKNHKWF ATTDWIAIYQ RKVEA PFIP KFKGPGDTSN FDDYEEEEIR V(SEP)INEKCGKE FTEF

UniProtKB: cAMP-dependent protein kinase catalytic subunit alpha

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Macromolecule #2: cAMP-dependent protein kinase type I-alpha regulatory subunit

MacromoleculeName: cAMP-dependent protein kinase type I-alpha regulatory subunit
type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Bos taurus (domestic cattle)
Molecular weightTheoretical: 47.41541 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MRGSHHHHHH GMASMTGGQQ MGRDLYDDDD KDPSSRSAAG TMASGTTASE EERSLRECEL YVQKHNIQAL LKDSIVQLCT ARPERPMAF LREYFEKLEK EEAKQIQNLQ KAGSRADSRE DEISPPPPNP VVKGRRRRGA ISAEVYTEED AASYVRKVIP K DYKTMAAL ...String:
MRGSHHHHHH GMASMTGGQQ MGRDLYDDDD KDPSSRSAAG TMASGTTASE EERSLRECEL YVQKHNIQAL LKDSIVQLCT ARPERPMAF LREYFEKLEK EEAKQIQNLQ KAGSRADSRE DEISPPPPNP VVKGRRRRGA ISAEVYTEED AASYVRKVIP K DYKTMAAL AKAIEKNVLF SHLDDNERSD IFDAMFPVSF IAGETVIQQG DEGDNFYVID QGEMDVYVNN EWATSVGEGG SF GELALIY GTPRAATVKA KTNVKLWGID RDSYRRILMG STLRKRKMYE EFLSKVSILE SLDKWERLTV ADALEPVQFE DGQ KIVVQG EPGDEFFIIL EGSAAVLQRR SENEEFVEVG RLGPSDYFGE IALLMNRPRA ATVVARGPLK CVKLDRPRFE RVLG PCSDI LKRNIQQYNS FVSLSV

UniProtKB: cAMP-dependent protein kinase type I-alpha regulatory subunit

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Macromolecule #3: ADENOSINE-5'-TRIPHOSPHATE

MacromoleculeName: ADENOSINE-5'-TRIPHOSPHATE / type: ligand / ID: 3 / Number of copies: 1 / Formula: ATP
Molecular weightTheoretical: 507.181 Da
Chemical component information

ChemComp-ATP:
ADENOSINE-5'-TRIPHOSPHATE / ATP, energy-carrying molecule*YM

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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

Concentration0.6 mg/mL
BufferpH: 7
Component:
ConcentrationFormulaName
50.0 mMMOPS3-(N-morpholino)propanesulfonic acid
50.0 mMNaClSodium Chloride
5.0 mMMgCl2Magnesium Chloride
0.1 mMATPAdenosine 5-Triphosphate
1.0 mMB-ME2-Mercaptoethanol
GridModel: Quantifoil R2/1 / Pretreatment - Type: GLOW DISCHARGE
VitrificationCryogen name: ETHANE / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TALOS ARCTICA
Image recordingFilm or detector model: FEI FALCON IV (4k x 4k) / Number real images: 4559 / Average electron dose: 49.66 e/Å2
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: SPOT SCAN / Imaging mode: OTHER / Nominal defocus max: 2.2 µm / Nominal defocus min: 1.0 µm
Experimental equipment
Model: Talos Arctica / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 2683140
Startup modelType of model: NONE
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 4.18 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 4.2) / Number images used: 95294
Initial angle assignmentType: NOT APPLICABLE
Final angle assignmentType: NOT APPLICABLE / Software - Name: cryoSPARC (ver. 4.2)
Final 3D classificationNumber classes: 3 / Software - Name: cryoSPARC (ver. 4.2)

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Atomic model buiding 1

Initial model
PDB IDChain

3fhi

source_name: PDB, initial_model_type: experimental model
source_name: ITasser, initial_model_type: in silico model
RefinementSpace: REAL / Protocol: RIGID BODY FIT / Overall B value: 369.72
Output model

PDB-9edc:
Reset Type-I Protein Kinase A Holoenzyme

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