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- PDB-6ynt: Crystal structure of the cAMP-dependent protein kinase A in compl... -

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Basic information

Entry
Database: PDB / ID: 6ynt
TitleCrystal structure of the cAMP-dependent protein kinase A in complex with aminofasudil and PKI (5-24)
Components(cAMP-dependent protein kinase ...) x 2
KeywordsTRANSFERASE / phosphotransferase / signalling pathways / glycogen metabolism / serine/threonine kinase
Function / homology
Function and homology information


negative regulation of cAMP/PKA signal transduction / cAMP-dependent protein kinase inhibitor activity / cAMP-dependent protein kinase / regulation of protein processing / cAMP-dependent protein kinase activity / protein localization to lipid droplet / cAMP-dependent protein kinase complex / regulation of bicellular tight junction assembly / cellular response to parathyroid hormone stimulus / negative regulation of protein import into nucleus ...negative regulation of cAMP/PKA signal transduction / cAMP-dependent protein kinase inhibitor activity / cAMP-dependent protein kinase / regulation of protein processing / cAMP-dependent protein kinase activity / protein localization to lipid droplet / cAMP-dependent protein kinase complex / regulation of bicellular tight junction assembly / cellular response to parathyroid hormone stimulus / negative regulation of protein import into nucleus / regulation of osteoblast differentiation / cellular response to cold / sperm capacitation / negative regulation of glycolytic process through fructose-6-phosphate / ciliary base / protein kinase A regulatory subunit binding / protein kinase A catalytic subunit binding / intracellular potassium ion homeostasis / mesoderm formation / plasma membrane raft / axoneme / sperm flagellum / postsynaptic modulation of chemical synaptic transmission / regulation of G2/M transition of mitotic cell cycle / negative regulation of TORC1 signaling / regulation of proteasomal protein catabolic process / positive regulation of gluconeogenesis / protein serine/threonine/tyrosine kinase activity / cellular response to glucagon stimulus / acrosomal vesicle / protein export from nucleus / positive regulation of phagocytosis / positive regulation of protein export from nucleus / negative regulation of smoothened signaling pathway / neural tube closure / neuromuscular junction / cellular response to glucose stimulus / positive regulation of cholesterol biosynthetic process / adenylate cyclase-inhibiting G protein-coupled receptor signaling pathway / positive regulation of insulin secretion / mRNA processing / adenylate cyclase-activating G protein-coupled receptor signaling pathway / manganese ion binding / cellular response to heat / postsynapse / regulation of cell cycle / nuclear speck / protein domain specific binding / protein serine kinase activity / protein serine/threonine kinase activity / ubiquitin protein ligase binding / centrosome / protein kinase binding / perinuclear region of cytoplasm / glutamatergic synapse / magnesium ion binding / negative regulation of transcription by RNA polymerase II / mitochondrion / ATP binding / nucleus / cytoplasm / cytosol
Similarity search - Function
cAMP-dependent protein kinase inhibitor / cAMP-dependent protein kinase inhibitor / cAMP-dependent protein kinase catalytic subunit / Extension to Ser/Thr-type protein kinases / AGC-kinase, C-terminal / AGC-kinase C-terminal domain profile. / Serine/threonine-protein kinase, active site / Serine/Threonine protein kinases active-site signature. / Protein kinase domain / Serine/Threonine protein kinases, catalytic domain ...cAMP-dependent protein kinase inhibitor / cAMP-dependent protein kinase inhibitor / cAMP-dependent protein kinase catalytic subunit / Extension to Ser/Thr-type protein kinases / AGC-kinase, C-terminal / AGC-kinase C-terminal domain profile. / 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
5-(1,4-diazepan-1-ylsulfonyl)isoquinolin-1-amine / cAMP-dependent protein kinase catalytic subunit alpha / cAMP-dependent protein kinase inhibitor alpha
Similarity search - Component
Biological speciesCricetulus griseus (Chinese hamster)
Mus musculus (house mouse)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 1.52 Å
AuthorsOebbeke, M. / Gerber, H.-D. / Heine, A. / Klebe, G.
CitationJournal: Chemmedchem / Year: 2021
Title: Two Methods, One Goal: Structural Differences between Cocrystallization and Crystal Soaking to Discover Ligand Binding Poses.
Authors: Wienen-Schmidt, B. / Oebbeke, M. / Ngo, K. / Heine, A. / Klebe, G.
History
DepositionApr 14, 2020Deposition site: PDBE / Processing site: PDBE
Revision 1.0Oct 14, 2020Provider: repository / Type: Initial release
Revision 1.1Oct 21, 2020Group: Database references / Category: citation / citation_author
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year
Revision 1.2Jan 20, 2021Group: Database references / Category: citation / citation_author
Item: _citation.journal_volume / _citation.page_first ..._citation.journal_volume / _citation.page_first / _citation.page_last / _citation.title / _citation.year / _citation_author.name
Revision 1.3Jan 24, 2024Group: Data collection / Database references / Refinement description
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / database_2 / pdbx_initial_refinement_model
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession
Revision 1.4Nov 13, 2024Group: Structure summary / Category: pdbx_entry_details / pdbx_modification_feature / Item: _pdbx_entry_details.has_protein_modification

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: cAMP-dependent protein kinase catalytic subunit alpha
B: cAMP-dependent protein kinase inhibitor alpha
hetero molecules


Theoretical massNumber of molelcules
Total (without water)44,1578
Polymers43,3402
Non-polymers8176
Water4,864270
1


  • Idetical with deposited unit
  • defined by author&software
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area3150 Å2
ΔGint-19 kcal/mol
Surface area15780 Å2
MethodPISA
Unit cell
Length a, b, c (Å)72.524, 74.460, 80.359
Angle α, β, γ (deg.)90.000, 90.000, 90.000
Int Tables number19
Space group name H-MP212121
Space group name HallP2ac2ab
Symmetry operation#1: x,y,z
#2: x+1/2,-y+1/2,-z
#3: -x,y+1/2,-z+1/2
#4: -x+1/2,-y,z+1/2

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Components

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CAMP-dependent protein kinase ... , 2 types, 2 molecules AB

#1: Protein cAMP-dependent protein kinase catalytic subunit alpha / PKA C-alpha


Mass: 41113.766 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Cricetulus griseus (Chinese hamster) / Gene: PRKACA / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: P25321, cAMP-dependent protein kinase
#2: Protein/peptide cAMP-dependent protein kinase inhibitor alpha / PKI-alpha / cAMP-dependent protein kinase inhibitor / muscle/brain isoform


Mass: 2226.411 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) Mus musculus (house mouse) / References: UniProt: P63248

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Non-polymers , 5 types, 276 molecules

#3: Chemical ChemComp-MPD / (4S)-2-METHYL-2,4-PENTANEDIOL


Mass: 118.174 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C6H14O2 / Comment: precipitant*YM
#4: Chemical ChemComp-DMS / DIMETHYL SULFOXIDE


Mass: 78.133 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C2H6OS / Comment: DMSO, precipitant*YM
#5: Chemical ChemComp-LRT / 5-(1,4-diazepan-1-ylsulfonyl)isoquinolin-1-amine


Mass: 306.383 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C14H18N4O2S / Feature type: SUBJECT OF INVESTIGATION
#6: Chemical ChemComp-MRD / (4R)-2-METHYLPENTANE-2,4-DIOL


Mass: 118.174 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C6H14O2 / Comment: precipitant*YM
#7: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 270 / Source method: isolated from a natural source / Formula: H2O

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Details

Has ligand of interestY
Has protein modificationY

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Experimental details

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Experiment

ExperimentMethod: X-RAY DIFFRACTION / Number of used crystals: 1

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Sample preparation

CrystalDensity Matthews: 2.64 Å3/Da / Density % sol: 53.49 %
Crystal growTemperature: 277 K / Method: vapor diffusion, hanging drop / pH: 6.9
Details: 0.2 mM PKA , 100 mM MBT (MES/Bis-Tris Puffer pH 6.9), 1 mM DTT, 0.1 mM EDTA, 75 mM LiCl, 0.2 mM Mega8, 0.5 mM PKI, 16-23% (v/v) Methanol. Soaking in buffer with 10% (v/v) of aminofasudil ...Details: 0.2 mM PKA , 100 mM MBT (MES/Bis-Tris Puffer pH 6.9), 1 mM DTT, 0.1 mM EDTA, 75 mM LiCl, 0.2 mM Mega8, 0.5 mM PKI, 16-23% (v/v) Methanol. Soaking in buffer with 10% (v/v) of aminofasudil (50mM stock) in DMSO and 30% (v/v) MPD

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Data collection

DiffractionMean temperature: 100 K / Serial crystal experiment: N
Diffraction sourceSource: SYNCHROTRON / Site: BESSY / Beamline: 14.1 / Wavelength: 0.9184 Å
DetectorType: DECTRIS PILATUS 6M / Detector: PIXEL / Date: Jul 11, 2019
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 0.9184 Å / Relative weight: 1
ReflectionResolution: 1.52→43.63 Å / Num. obs: 67316 / % possible obs: 99.5 % / Redundancy: 4.4 % / Biso Wilson estimate: 20.29 Å2 / Rsym value: 0.044 / Net I/σ(I): 17.08
Reflection shellResolution: 1.52→1.61 Å / Mean I/σ(I) obs: 2.97 / Num. unique obs: 10749 / Rsym value: 0.44 / % possible all: 99.6

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Processing

Software
NameVersionClassification
PHASERphasing
PHENIX1.17.1_3660refinement
XDSdata reduction
XDSdata scaling
Cootmodel building
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: 6F14

6f14


Resolution: 1.52→43.63 Å / SU ML: 0.131 / Cross valid method: FREE R-VALUE / σ(F): 1.37 / Phase error: 17.1436
RfactorNum. reflection% reflection
Rfree0.1914 3366 5 %
Rwork0.1694 --
obs0.1705 67315 99.55 %
Solvent computationShrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å
Displacement parametersBiso mean: 26.59 Å2
Refinement stepCycle: LAST / Resolution: 1.52→43.63 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms2881 0 53 270 3204
Refine LS restraints
Refine-IDTypeDev idealNumber
X-RAY DIFFRACTIONf_bond_d0.00763093
X-RAY DIFFRACTIONf_angle_d0.97144214
X-RAY DIFFRACTIONf_chiral_restr0.0555451
X-RAY DIFFRACTIONf_plane_restr0.0071550
X-RAY DIFFRACTIONf_dihedral_angle_d21.77671118
LS refinement shell
Resolution (Å)Rfactor RfreeNum. reflection RfreeRfactor RworkNum. reflection RworkRefine-ID% reflection obs (%)
1.52-1.540.22521370.20782603X-RAY DIFFRACTION98.92
1.54-1.560.27891390.20512642X-RAY DIFFRACTION99.93
1.56-1.590.21390.19752635X-RAY DIFFRACTION99.82
1.59-1.610.22891390.19822650X-RAY DIFFRACTION99.75
1.61-1.640.2331390.18522643X-RAY DIFFRACTION99.78
1.64-1.670.2171380.18682614X-RAY DIFFRACTION99.53
1.67-1.70.21311390.18682655X-RAY DIFFRACTION99.86
1.7-1.740.24861400.17552645X-RAY DIFFRACTION99.93
1.74-1.780.21151390.17172649X-RAY DIFFRACTION99.86
1.78-1.820.19141390.16132648X-RAY DIFFRACTION99.54
1.82-1.860.20381400.16032644X-RAY DIFFRACTION99.86
1.86-1.910.18741390.15042657X-RAY DIFFRACTION99.89
1.91-1.970.1861390.16252636X-RAY DIFFRACTION99.36
1.97-2.030.17821390.16242638X-RAY DIFFRACTION99.32
2.03-2.110.16521400.1572662X-RAY DIFFRACTION99.93
2.11-2.190.16541420.15872687X-RAY DIFFRACTION99.82
2.19-2.290.2031390.15592643X-RAY DIFFRACTION99.86
2.29-2.410.1851400.15472676X-RAY DIFFRACTION99.29
2.41-2.560.18531410.16082664X-RAY DIFFRACTION99.01
2.56-2.760.21471410.172690X-RAY DIFFRACTION99.68
2.76-3.040.1861420.17262690X-RAY DIFFRACTION99.68
3.04-3.480.19141430.17532717X-RAY DIFFRACTION99.27
3.48-4.380.18071430.15632718X-RAY DIFFRACTION98.86
4.38-43.630.18521500.18792843X-RAY DIFFRACTION98.65
Refinement TLS params.

Method: refined / Refine-ID: X-RAY DIFFRACTION

IDL11 (°2)L12 (°2)L13 (°2)L22 (°2)L23 (°2)L33 (°2)S11 (Å °)S12 (Å °)S13 (Å °)S21 (Å °)S22 (Å °)S23 (Å °)S31 (Å °)S32 (Å °)S33 (Å °)T11 (Å2)T12 (Å2)T13 (Å2)T22 (Å2)T23 (Å2)T33 (Å2)Origin x (Å)Origin y (Å)Origin z (Å)
10.7796057611380.07699767311170.09158440348991.27921466276-0.04795810914061.33525735266-0.0376369650830.0237926287088-0.0248196109280.03919622951850.05314131912880.0975576251767-0.0230231520139-0.135714929848-0.01546002575280.0940781346573-0.0104625396750.01350600619080.2073589767820.003217681147650.182897023737-32.9434359548-11.00176383877.3155137704
20.5636806000960.248213346467-0.1088660554721.36088326135-0.9444282334421.60685015636-0.03346098338090.05218737958280.016474892430.02847849750790.04608234698190.0765569601653-0.205388935993-0.126643575405-0.02125700187340.1303867304710.010228121151-0.005436326445930.171525866524-0.005001495412940.155396456347-28.69392327940.2383616180340.933448362172
31.203310074820.07061871276750.1142134486471.14798901311-0.3521373923742.165113420990.0340092807893-0.03528636966310.2040965504050.169413740018-0.0981421811676-0.101837256116-0.6267972023230.2432389999760.03913809900550.31731076189-0.0721430877274-0.01928493584350.1626619370470.008969996156090.179969293615-18.510437512511.96421739831.97728347015
40.4022112024910.618953235256-0.05268853979160.949821653779-0.07700630010321.56392039929-0.05806685118980.164691525582-0.0138347652254-0.191942950260.07924729646050.08236435422390.00334923356655-0.2280301939430.01535012699190.115596789770.01800219863190.005483803351210.2405070989870.009708659936810.203078689489-32.0801034123-9.15675030362-2.11908722976
52.99502340793-2.927783419951.028214171917.67195312246-2.095958426845.369081665770.00331412230162-0.07799585639180.02683912015490.00646087177555-0.1546639606640.255257016665-0.151980475847-0.03674634638430.1435982680430.170845095156-0.0472609447085-0.006143834683190.2166977614040.001418341857120.157079654531-13.21293272397.79386701541-16.3059530456
62.627244809331.5863177452-4.489381697686.74916178041-3.250462390448.47892370289-0.302622738486-0.013081354526-0.268614110225-0.0883856265532-0.150901213056-0.167164108890.3076179123770.3254420258360.4753539192060.153649489911-0.01004586672460.02674071514360.1689428413160.01179097369170.184020769325-14.5265421072-3.19682688074-5.66815843233
Refinement TLS group
IDRefine-IDRefine TLS-IDSelection details
1X-RAY DIFFRACTION1chain 'A' and (resid 5 through 114 )
2X-RAY DIFFRACTION2chain 'A' and (resid 115 through 198 )
3X-RAY DIFFRACTION3chain 'A' and (resid 199 through 297 )
4X-RAY DIFFRACTION4chain 'A' and (resid 298 through 350 )
5X-RAY DIFFRACTION5chain 'B' and (resid 5 through 12 )
6X-RAY DIFFRACTION6chain 'B' and (resid 13 through 23 )

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