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- PDB-8bik: Crystal structure of human AMPK heterotrimer in complex with allo... -

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Basic information

Entry
Database: PDB / ID: 8bik
TitleCrystal structure of human AMPK heterotrimer in complex with allosteric activator C455
Components
  • (5'-AMP-activated protein kinase subunit ...) x 2
  • 5'-AMP-activated protein kinase catalytic subunit alpha-2
KeywordsSIGNALING PROTEIN / AMPK activator / allosteric activator / ADaM site / structure-based drug design / selectivity
Function / homology
Function and homology information


nail development / [hydroxymethylglutaryl-CoA reductase (NADPH)] kinase / [hydroxymethylglutaryl-CoA reductase (NADPH)] kinase activity / regulation of stress granule assembly / AMPK inhibits chREBP transcriptional activation activity / histone H2BS36 kinase activity / cAMP-dependent protein kinase regulator activity / AMP-activated protein kinase activity / lipid droplet disassembly / Lipophagy ...nail development / [hydroxymethylglutaryl-CoA reductase (NADPH)] kinase / [hydroxymethylglutaryl-CoA reductase (NADPH)] kinase activity / regulation of stress granule assembly / AMPK inhibits chREBP transcriptional activation activity / histone H2BS36 kinase activity / cAMP-dependent protein kinase regulator activity / AMP-activated protein kinase activity / lipid droplet disassembly / Lipophagy / regulation of carbon utilization / import into nucleus / nucleotide-activated protein kinase complex / negative regulation of hepatocyte apoptotic process / Energy dependent regulation of mTOR by LKB1-AMPK / Carnitine shuttle / protein kinase regulator activity / negative regulation of TOR signaling / Activation of PPARGC1A (PGC-1alpha) by phosphorylation / Nuclear events mediated by NFE2L2 / regulation of glycolytic process / cAMP-dependent protein kinase activity / protein localization to lipid droplet / negative regulation of tubulin deacetylation / AMP binding / Macroautophagy / cholesterol biosynthetic process / lipid biosynthetic process / response to muscle activity / positive regulation of protein kinase activity / positive regulation of macroautophagy / regulation of macroautophagy / fatty acid homeostasis / cellular response to nutrient levels / cellular response to glucose starvation / energy homeostasis / Activation of AMPK downstream of NMDARs / positive regulation of protein localization / negative regulation of TORC1 signaling / positive regulation of autophagy / protein serine/threonine/tyrosine kinase activity / positive regulation of gluconeogenesis / cellular response to calcium ion / regulation of microtubule cytoskeleton organization / positive regulation of glycolytic process / TP53 Regulates Metabolic Genes / Translocation of SLC2A4 (GLUT4) to the plasma membrane / cellular response to glucose stimulus / regulation of circadian rhythm / ADP binding / autophagy / cellular response to xenobiotic stimulus / Wnt signaling pathway / cytoplasmic stress granule / fatty acid biosynthetic process / rhythmic process / glucose homeostasis / cellular response to prostaglandin E stimulus / positive regulation of cold-induced thermogenesis / cellular response to oxidative stress / spermatogenesis / Regulation of TP53 Activity through Phosphorylation / protein phosphorylation / protein kinase activity / non-specific serine/threonine protein kinase / regulation of cell cycle / nuclear speck / ciliary basal body / axon / negative regulation of gene expression / protein serine kinase activity / neuronal cell body / protein serine/threonine kinase activity / dendrite / chromatin binding / protein kinase binding / negative regulation of apoptotic process / protein-containing complex binding / Golgi apparatus / signal transduction / nucleoplasm / ATP binding / metal ion binding / nucleus / membrane / cytosol / cytoplasm
Similarity search - Function
PRKAA2, UBA-like autoinhibitory domain / 5'-AMP-activated protein kinase catalytic subunit alpha-2, C-terminal / : / AMP-activated protein kinase, alpha subunit, autoinhibitory domain / : / Association with the SNF1 complex (ASC) domain / ASC domain superfamily / : / 5'-AMP-activated protein kinase beta subunit, interaction domain / 5'-AMP-activated protein kinase beta subunit, interation domain ...PRKAA2, UBA-like autoinhibitory domain / 5'-AMP-activated protein kinase catalytic subunit alpha-2, C-terminal / : / AMP-activated protein kinase, alpha subunit, autoinhibitory domain / : / Association with the SNF1 complex (ASC) domain / ASC domain superfamily / : / 5'-AMP-activated protein kinase beta subunit, interaction domain / 5'-AMP-activated protein kinase beta subunit, interation domain / AMPK, C-terminal adenylate sensor domain / Adenylate sensor of SNF1-like protein kinase / AMP-activated protein kinase, glycogen-binding domain / Glycogen recognition site of AMP-activated protein kinase / KA1 domain/Ssp2, C-terminal / Domain in cystathionine beta-synthase and other proteins. / CBS domain superfamily / CBS domain / CBS domain / CBS domain profile. / Immunoglobulin E-set / 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. / Immunoglobulin-like fold / Protein kinase domain profile. / Protein kinase domain / Protein kinase-like domain superfamily
Similarity search - Domain/homology
ADENOSINE MONOPHOSPHATE / : / STAUROSPORINE / 5'-AMP-activated protein kinase subunit gamma-1 / 5'-AMP-activated protein kinase catalytic subunit alpha-2 / 5'-AMP-activated protein kinase subunit beta-1
Similarity search - Component
Biological speciesHomo sapiens (human)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / molecular replacement / Resolution: 2.5 Å
AuthorsSchimpl, M. / Mather, K.M. / Boland, M.L. / Rivers, E.L. / Srivastava, A. / Hemsley, P. / Robinson, J. / Wan, P.T. / Hansen, J. / Read, J.A. ...Schimpl, M. / Mather, K.M. / Boland, M.L. / Rivers, E.L. / Srivastava, A. / Hemsley, P. / Robinson, J. / Wan, P.T. / Hansen, J. / Read, J.A. / Trevaskis, J.L. / Smith, D.M.
Funding support1items
OrganizationGrant numberCountry
Not funded
CitationJournal: Biorxiv / Year: 2024
Title: Direct beta 1/ beta 2 AMPK activation reduces liver steatosis but not fibrosis in a mouse model of non-alcoholic steatohepatitis
Authors: Mather, K.M. / Boland, M.L. / Rivers, E.L. / Srivastava, A. / Schimpl, M. / Hemsley, P. / Robinson, J. / Wan, P. / Hansen, J. / Read, J.A. / Trevaskis, J.L. / Smith, D.M.
History
DepositionNov 2, 2022Deposition site: PDBE / Processing site: PDBE
Revision 1.0May 15, 2024Provider: repository / Type: Initial release
Revision 1.1Jun 12, 2024Group: 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.title / _citation.year
Revision 1.2Oct 16, 2024Group: Structure summary / Category: pdbx_entry_details / pdbx_modification_feature

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: 5'-AMP-activated protein kinase catalytic subunit alpha-2
B: 5'-AMP-activated protein kinase subunit beta-1
C: 5'-AMP-activated protein kinase subunit gamma-1
D: 5'-AMP-activated protein kinase catalytic subunit alpha-2
E: 5'-AMP-activated protein kinase subunit beta-1
F: 5'-AMP-activated protein kinase subunit gamma-1
hetero molecules


Theoretical massNumber of molelcules
Total (without water)266,75415
Polymers262,9996
Non-polymers3,7559
Water8,683482
1
A: 5'-AMP-activated protein kinase catalytic subunit alpha-2
B: 5'-AMP-activated protein kinase subunit beta-1
C: 5'-AMP-activated protein kinase subunit gamma-1
hetero molecules


Theoretical massNumber of molelcules
Total (without water)133,2037
Polymers131,4993
Non-polymers1,7044
Water543
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area16020 Å2
ΔGint-98 kcal/mol
Surface area41640 Å2
MethodPISA
2
D: 5'-AMP-activated protein kinase catalytic subunit alpha-2
E: 5'-AMP-activated protein kinase subunit beta-1
F: 5'-AMP-activated protein kinase subunit gamma-1
hetero molecules


Theoretical massNumber of molelcules
Total (without water)133,5518
Polymers131,4993
Non-polymers2,0515
Water543
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area16450 Å2
ΔGint-100 kcal/mol
Surface area40380 Å2
MethodPISA
Unit cell
Length a, b, c (Å)75.313, 127.975, 139.215
Angle α, β, γ (deg.)90.000, 92.910, 90.000
Int Tables number4
Space group name H-MP1211

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Components

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Protein , 1 types, 2 molecules AD

#1: Protein 5'-AMP-activated protein kinase catalytic subunit alpha-2 / AMPK subunit alpha-2 / Acetyl-CoA carboxylase kinase / ACACA kinase / Hydroxymethylglutaryl-CoA ...AMPK subunit alpha-2 / Acetyl-CoA carboxylase kinase / ACACA kinase / Hydroxymethylglutaryl-CoA reductase kinase / HMGCR kinase


Mass: 63368.684 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: PRKAA2, AMPK, AMPK2 / Production host: Escherichia coli BL21(DE3) (bacteria)
References: UniProt: P54646, non-specific serine/threonine protein kinase, EC: 2.7.11.27, [hydroxymethylglutaryl-CoA reductase (NADPH)] kinase

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5'-AMP-activated protein kinase subunit ... , 2 types, 4 molecules BECF

#2: Protein 5'-AMP-activated protein kinase subunit beta-1 / AMPK subunit beta-1 / AMPKb


Mass: 30504.299 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: PRKAB1, AMPK / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: Q9Y478
#3: Protein 5'-AMP-activated protein kinase subunit gamma-1 / AMPK gamma1 / AMPK subunit gamma-1 / AMPKg


Mass: 37626.289 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: PRKAG1 / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: P54619

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Non-polymers , 4 types, 491 molecules

#4: Chemical ChemComp-STU / STAUROSPORINE


Mass: 466.531 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C28H26N4O3 / Comment: antibiotic*YM
#5: Chemical ChemComp-QTN / (3~{R},3~{a}~{R},6~{R},6~{a}~{R})-6-[[6-chloranyl-5-[4-[4-[[dimethyl(oxidanyl)-$l^{4}-sulfanyl]amino]phenyl]phenyl]-3~{H}-imidazo[4,5-b]pyridin-2-yl]oxy]-2,3,3~{a},5,6,6~{a}-hexahydrofuro[3,2-b]furan-3-ol


Mass: 543.034 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C26H27ClN4O5S
#6: Chemical
ChemComp-AMP / ADENOSINE MONOPHOSPHATE


Mass: 347.221 Da / Num. of mol.: 5 / Source method: obtained synthetically / Formula: C10H14N5O7P / Comment: AMP*YM
#7: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 482 / Source method: isolated from a natural source / Formula: H2O

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Details

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: 2.55 Å3/Da / Density % sol: 51.72 %
Crystal growTemperature: 293 K / Method: vapor diffusion / pH: 7.2
Details: AMPK a2b1g1 (5 mg/ml: 38 uM) was combined with AMP (4-fold excess), staurosporine (1.2-fold excess) and activator 455 (3-fold excess): and crystallised at 20 C by mixing 2 ul of the protein ...Details: AMPK a2b1g1 (5 mg/ml: 38 uM) was combined with AMP (4-fold excess), staurosporine (1.2-fold excess) and activator 455 (3-fold excess): and crystallised at 20 C by mixing 2 ul of the protein solution with 1 ul of 8 % PEG3350, 0.3 M guanidine hydrochloride, 0.1 M PIPES buffer pH 7.2.

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Data collection

DiffractionMean temperature: 100 K / Serial crystal experiment: N
Diffraction sourceSource: SYNCHROTRON / Site: Diamond / Beamline: I02 / Wavelength: 0.97949 Å
DetectorType: DECTRIS PILATUS 6M-F / Detector: PIXEL / Date: Apr 29, 2016
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 0.97949 Å / Relative weight: 1
ReflectionResolution: 2.5→75.22 Å / Num. obs: 91016 / % possible obs: 99.9 % / Redundancy: 3.3 % / Biso Wilson estimate: 69.44 Å2 / CC1/2: 0.988 / Rmerge(I) obs: 0.069 / Rpim(I) all: 0.045 / Rrim(I) all: 0.082 / Net I/σ(I): 8.9 / Num. measured all: 300129 / Scaling rejects: 73
Reflection shell

Diffraction-ID: 1

Resolution (Å)Redundancy (%)Rmerge(I) obsNum. measured allNum. unique obsCC1/2Rpim(I) allRrim(I) allNet I/σ(I) obs% possible all
2.5-2.563.40.8632243866890.5460.5521.0271.499.9
11.18-75.223.10.04329510710.9380.0290.052699.7

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Phasing

PhasingMethod: molecular replacement

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Processing

Software
NameVersionClassification
Aimless0.5.23data scaling
PHASERphasing
BUSTER2.11.7refinement
PDB_EXTRACT3.25data extraction
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: pdbid 4CFF

4cff


Resolution: 2.5→69.52 Å / Cor.coef. Fo:Fc: 0.955 / Cor.coef. Fo:Fc free: 0.941 / SU R Cruickshank DPI: 0.336 / Cross valid method: THROUGHOUT / σ(F): 0 / SU R Blow DPI: 0.32 / SU Rfree Blow DPI: 0.217 / SU Rfree Cruickshank DPI: 0.223
RfactorNum. reflection% reflectionSelection details
Rfree0.215 4335 4.81 %RANDOM
Rwork0.181 ---
obs0.183 90076 98.9 %-
Displacement parametersBiso max: 238.29 Å2 / Biso mean: 67.72 Å2 / Biso min: 22.6 Å2
Baniso -1Baniso -2Baniso -3
1--2.5921 Å20 Å2-0.3767 Å2
2---0.0511 Å20 Å2
3---2.6433 Å2
Refine analyzeLuzzati coordinate error obs: 0.31 Å
Refinement stepCycle: final / Resolution: 2.5→69.52 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms14741 0 259 482 15482
Biso mean--58.42 57.97 -
Num. residues----1846
Refine LS restraints
Refine-IDTypeNumberRestraint functionWeightDev ideal
X-RAY DIFFRACTIONt_dihedral_angle_d5284SINUSOIDAL2
X-RAY DIFFRACTIONt_trig_c_planes
X-RAY DIFFRACTIONt_gen_planes2566HARMONIC5
X-RAY DIFFRACTIONt_it15380HARMONIC20
X-RAY DIFFRACTIONt_nbd
X-RAY DIFFRACTIONt_improper_torsion
X-RAY DIFFRACTIONt_pseud_angle
X-RAY DIFFRACTIONt_chiral_improper_torsion2004SEMIHARMONIC5
X-RAY DIFFRACTIONt_sum_occupancies
X-RAY DIFFRACTIONt_utility_distance
X-RAY DIFFRACTIONt_utility_angle
X-RAY DIFFRACTIONt_utility_torsion
X-RAY DIFFRACTIONt_ideal_dist_contact16818SEMIHARMONIC4
X-RAY DIFFRACTIONt_bond_d15380HARMONIC20.01
X-RAY DIFFRACTIONt_angle_deg20916HARMONIC21.13
X-RAY DIFFRACTIONt_omega_torsion3.04
X-RAY DIFFRACTIONt_other_torsion20.1
LS refinement shellResolution: 2.5→2.52 Å / Rfactor Rfree error: 0 / Total num. of bins used: 50
RfactorNum. reflection% reflection
Rfree0.3008 83 4.61 %
Rwork0.2375 1719 -
all0.2404 1802 -
obs--95.41 %
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)
10.6543-0.43130.05560.8766-0.27380.7545-0.1299-0.15790.25090.32550.1458-0.0826-0.5608-0.1982-0.01590.28630.1479-0.0243-0.1258-0.071-0.0449-2.57828.539521.427
20.2551-0.62430.13211.1068-0.6580.3783-0.1503-0.10290.16710.54920.1159-0.2872-0.3714-0.0540.03440.28760.0445-0.1199-0.0802-0.0372-0.0359-0.033246.530318.7924
30.9165-0.2730.05661.04240.18881.5727-0.0835-0.08340.19140.0970.0472-0.3856-0.44340.32610.03630.2857-0.1077-0.12360.1813-0.01950.13831.72816.582335.2288
40.67070.7425-0.40290.9835-0.18710.65930.11350.23210.3810.0090.08060.3378-0.1005-0.1888-0.19420.03630.05520.03120.23470.11240.379834.01470.922845.2556
50.6006-0.1188-0.708100.66650.926-0.13710.16060.0101-0.35450.1092-0.18620.3342-0.08990.0279-0.06470.0475-0.08950.32110.13230.173336.534454.294751.5562
60.56430.7933-0.30472.88960.13890.69570.2538-0.27980.39210.08960.00930.2854-0.50340.1894-0.26310.1603-0.08180.27480.0165-0.1580.124568.836495.646335.0758
Refinement TLS group
IDRefine-IDRefine TLS-IDSelection detailsAuth asym-IDAuth seq-ID
1X-RAY DIFFRACTION1{ A|* }A8 - 551
2X-RAY DIFFRACTION2{ B|* }B77 - 270
3X-RAY DIFFRACTION3{ C|* }C25 - 324
4X-RAY DIFFRACTION4{ D|* }D9 - 551
5X-RAY DIFFRACTION5{ E|* }E78 - 270
6X-RAY DIFFRACTION6{ F|* }F26 - 324

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