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- EMDB-18396: Cryo-EM structure of C-terminally truncated Apoptosis signal-regu... -

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Basic information

Entry
Database: EMDB / ID: EMD-18396
TitleCryo-EM structure of C-terminally truncated Apoptosis signal-regulating kinase 1 (ASK1)
Map datamain map, sharpened in phenix.autosharpen
Sample
  • Complex: C-terminally truncated Apoptosis signal-regulating kinase 1
    • Protein or peptide: Mitogen-activated protein kinase kinase kinase 5
KeywordsASK1 / MAP3K / MAPK signaling / thioredoxin / APOPTOSIS
Function / homology
Function and homology information


cellular response to reactive nitrogen species / neuron intrinsic apoptotic signaling pathway in response to oxidative stress / IRE1-TRAF2-ASK1 complex / protein kinase complex / mitogen-activated protein kinase kinase kinase / programmed necrotic cell death / JUN kinase kinase kinase activity / endothelial cell apoptotic process / positive regulation of p38MAPK cascade / intrinsic apoptotic signaling pathway in response to oxidative stress ...cellular response to reactive nitrogen species / neuron intrinsic apoptotic signaling pathway in response to oxidative stress / IRE1-TRAF2-ASK1 complex / protein kinase complex / mitogen-activated protein kinase kinase kinase / programmed necrotic cell death / JUN kinase kinase kinase activity / endothelial cell apoptotic process / positive regulation of p38MAPK cascade / intrinsic apoptotic signaling pathway in response to oxidative stress / positive regulation of cardiac muscle cell apoptotic process / p38MAPK cascade / positive regulation of cysteine-type endopeptidase activity involved in apoptotic process / MAP kinase kinase kinase activity / intrinsic apoptotic signaling pathway in response to endoplasmic reticulum stress / positive regulation of protein kinase activity / positive regulation of myoblast differentiation / stress-activated MAPK cascade / positive regulation of JUN kinase activity / JNK cascade / positive regulation of vascular associated smooth muscle cell proliferation / cellular response to amino acid starvation / response to endoplasmic reticulum stress / response to ischemia / apoptotic signaling pathway / positive regulation of JNK cascade / cellular response to hydrogen peroxide / MAPK cascade / cellular senescence / cellular response to tumor necrosis factor / protein phosphatase binding / neuron apoptotic process / Oxidative Stress Induced Senescence / protein kinase activity / positive regulation of apoptotic process / protein domain specific binding / external side of plasma membrane / protein phosphorylation / protein serine kinase activity / innate immune response / protein serine/threonine kinase activity / protein kinase binding / positive regulation of DNA-templated transcription / magnesium ion binding / protein homodimerization activity / protein-containing complex / ATP binding / identical protein binding / cytosol / cytoplasm
Similarity search - Function
MAP3K, deoxyribohydrolase domain / MAP3K, HisK-N-like globin domain / HisK-N-like globin domain of the ASK signalosome / Deoxyribohydrolase (DRHyd) domain of the ASK signalosome / MAP3K, TRAFs-binding domain / MAP3K, PH domain / MAP3K TRAFs-binding domain / ASK kinase PH domain / Sterile alpha motif/pointed domain superfamily / Serine/threonine-protein kinase, active site ...MAP3K, deoxyribohydrolase domain / MAP3K, HisK-N-like globin domain / HisK-N-like globin domain of the ASK signalosome / Deoxyribohydrolase (DRHyd) domain of the ASK signalosome / MAP3K, TRAFs-binding domain / MAP3K, PH domain / MAP3K TRAFs-binding domain / ASK kinase PH domain / Sterile alpha motif/pointed 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
Mitogen-activated protein kinase kinase kinase 5
Similarity search - Component
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.71 Å
AuthorsKosek D / Honzejkova K / Obsilova V / Obsil T
Funding support Czech Republic, 2 items
OrganizationGrant numberCountry
Czech Science Foundation19-00121S Czech Republic
Czech Academy of Sciences67985823 Czech Republic
CitationJournal: Elife / Year: 2024
Title: The cryo-EM structure of ASK1 reveals an asymmetric architecture allosterically modulated by TRX1.
Authors: Karolina Honzejkova / Dalibor Kosek / Veronika Obsilova / Tomas Obsil /
Abstract: Apoptosis signal-regulating kinase 1 (ASK1) is a crucial stress sensor, directing cells toward apoptosis, differentiation, and senescence via the p38 and JNK signaling pathways. ASK1 dysregulation ...Apoptosis signal-regulating kinase 1 (ASK1) is a crucial stress sensor, directing cells toward apoptosis, differentiation, and senescence via the p38 and JNK signaling pathways. ASK1 dysregulation has been associated with cancer and inflammatory, cardiovascular, and neurodegenerative diseases, among others. However, our limited knowledge of the underlying structural mechanism of ASK1 regulation hampers our ability to target this member of the MAP3K protein family towards developing therapeutic interventions for these disorders. Nevertheless, as a multidomain Ser/Thr protein kinase, ASK1 is regulated by a complex mechanism involving dimerization and interactions with several other proteins, including thioredoxin 1 (TRX1). Thus, the present study aims at structurally characterizing ASK1 and its complex with TRX1 using several biophysical techniques. As shown by cryo-EM analysis, in a state close to its active form, ASK1 is a compact and asymmetric dimer, which enables extensive interdomain and interchain interactions. These interactions stabilize the active conformation of the ASK1 kinase domain. In turn, TRX1 functions as a negative allosteric effector of ASK1, modifying the structure of the TRX1-binding domain and changing its interaction with the tetratricopeptide repeats domain. Consequently, TRX1 reduces access to the activation segment of the kinase domain. Overall, our findings not only clarify the role of ASK1 dimerization and inter-domain contacts but also provide key mechanistic insights into its regulation, thereby highlighting the potential of ASK1 protein-protein interactions as targets for anti-inflammatory therapy.
History
DepositionSep 6, 2023-
Header (metadata) releaseApr 3, 2024-
Map releaseApr 3, 2024-
UpdateApr 3, 2024-
Current statusApr 3, 2024Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_18396.png

Downloads & links

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Map

FileDownload / File: emd_18396.map.gz / Format: CCP4 / Size: 149.9 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotationmain map, sharpened in phenix.autosharpen
Voxel sizeX=Y=Z: 0.8336 Å
Density
Contour LevelBy AUTHOR: 3.0
Minimum - Maximum-26.769579 - 46.689900000000002
Average (Standard dev.)-0.000000000004257 (±1.0)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderZYX
Origin000
Dimensions340340340
Spacing340340340
CellA=B=C: 283.42398 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_18396_msk_1.map
Projections & Slices
AxesZYX

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Additional map: mask

Fileemd_18396_additional_1.map
Annotationmask
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Half map: halfmapA

Fileemd_18396_half_map_1.map
AnnotationhalfmapA
Projections & Slices
AxesZYX

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Half map: halfmapB

Fileemd_18396_half_map_2.map
AnnotationhalfmapB
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Sample components

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Entire : C-terminally truncated Apoptosis signal-regulating kinase 1

EntireName: C-terminally truncated Apoptosis signal-regulating kinase 1
Components
  • Complex: C-terminally truncated Apoptosis signal-regulating kinase 1
    • Protein or peptide: Mitogen-activated protein kinase kinase kinase 5

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Supramolecule #1: C-terminally truncated Apoptosis signal-regulating kinase 1

SupramoleculeName: C-terminally truncated Apoptosis signal-regulating kinase 1
type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 203.5 KDa

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Macromolecule #1: Mitogen-activated protein kinase kinase kinase 5

MacromoleculeName: Mitogen-activated protein kinase kinase kinase 5 / type: protein_or_peptide / ID: 1 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 101.908625 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MSRRTTVAYV INEASQGQLV VAESEALQSL REACETVGAT LETLHFGKLD FGETTVLDRF YNADIAVVEM SDAFRQPSLF YHLGVRESF SMANNIILYC DTNSDSLQSL KEIICQKNTM CTGNYTFVPY MITPHNKVYC CDSSFMKGLT ELMQPNFELL L GPICLPLV ...String:
MSRRTTVAYV INEASQGQLV VAESEALQSL REACETVGAT LETLHFGKLD FGETTVLDRF YNADIAVVEM SDAFRQPSLF YHLGVRESF SMANNIILYC DTNSDSLQSL KEIICQKNTM CTGNYTFVPY MITPHNKVYC CDSSFMKGLT ELMQPNFELL L GPICLPLV DRFIQLLKVA QASSSQYFRE SILNDIRKAR NLYTGKELAA ELARIRQRVD NIEVLTADIV INLLLSYRDI QD YDSIVKL VETLEKLPTF DLASHHHVKF HYAFALNRRN LPGDRAKALD IMIPMVQSEG QVASDMYCLV GRIYKDMFLD SNF TDTESR DHGASWFKKA FESEPTLQSG INYAVLLLAA GHQFESSFEL RKVGVKLSSL LGKKGNLEKL QSYWEVGFFL GASV LANDH MRVIQASEKL FKLKTPAWYL KSIVETILIY KHFVKLTTEQ PVAKQELVDF WMDFLVEATK TDVTVVRFPV LILEP TKIY QPSYLSINNE VEEKTISIWH VLPDDKKGIH EWNFSASSVR GVSISKFEER CCFLYVLHNS DDFQIYFCTE LHCKKF FEM VNTITEEKGR STEEGDCESD LLEYDYEYDE NGDRVVLGKG TYGIVYAGRD LSNQVRIAIK EIPERDSRYS QPLHEEI AL HKHLKHKNIV QYLGSFSENG FIKIFMEQVP GGSLSALLRS KWGPLKDNEQ TIGFYTKQIL EGLKYLHDNQ IVHRDIKG D NVLINTYSGV LKISDFGTSK RLAGINPCTE TFTGTLQYMA PEIIDKGPRG YGKAADIWSL GCTIIEMATG KPPFYELGE PQAAMFKVGM FKVHPEIPES MSAEAKAFIL KCFEPDPDKR ACANDLLVDE FLKVSSKKKK TQPKLSALSA GSNEYLRRIS LPVPVLVNS SHHHHHH

UniProtKB: Mitogen-activated protein kinase kinase kinase 5

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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

Concentration0.7 mg/mL
BufferpH: 7.5
Details: 20 mM Tris-HCl 7.5 150 mM NaCl 2 mM 2-mercaptoethanol 3.8 mM CHAPSO
GridModel: UltrAuFoil R1.2/1.3 / Material: GOLD / Mesh: 300 / Pretreatment - Type: PLASMA CLEANING / Pretreatment - Time: 45 sec. / Pretreatment - Atmosphere: OTHER / Details: Gatan Solarus II 955
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 298 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: OTHER / Imaging mode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 2.8000000000000003 µm / Nominal defocus min: 0.7000000000000001 µm / Nominal magnification: 105000
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Number grids imaged: 2 / Number real images: 5781 / Average electron dose: 40.0 e/Å2
Details: 40 frames 2704 micrographs with 0 degree tilt, 8691 micrographs with 40 degree tilt, 5781 selected for analysis
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 1126189
Startup modelType of model: NONE
Initial angle assignmentType: ANGULAR RECONSTITUTION / Software - Name: cryoSPARC (ver. 4.1.2)
Final 3D classificationSoftware - Name: cryoSPARC (ver. 4.1.2)
Final angle assignmentType: ANGULAR RECONSTITUTION / Software - Name: cryoSPARC (ver. 4.1.2)
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.71 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 4.1.2)
Details: Sharpening was done using phenix.autosharpen (1.19.1-4122)
Number images used: 131110
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial model
PDB IDChainDetails

2clq

chain_id: A, source_name: PDB, initial_model_type: experimental modelkinase domain

5ulm

chain_id: A, source_name: PDB, initial_model_type: experimental modelCRR

3-f1

residue_range: 94-271, source_name: AlphaFold, initial_model_type: in silico modelTBD initial model
RefinementSpace: REAL / Protocol: FLEXIBLE FIT
Output model

PDB-8qgy:
Cryo-EM structure of C-terminally truncated Apoptosis signal-regulating kinase 1 (ASK1)

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