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- PDB-9ggm: Cryo-EM structure of KBTBD4 P313PRR mutant-HDAC2 2:2 complex -

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Basic information

Entry
Database: PDB / ID: 9ggm
TitleCryo-EM structure of KBTBD4 P313PRR mutant-HDAC2 2:2 complex
Components
  • Histone deacetylase 2
  • Isoform 1 of Kelch repeat and BTB domain-containing protein 4
KeywordsLIGASE / E3 ligase / deacetylase / transcription / cancer mutation / protein complex
Function / homology
Function and homology information


positive regulation of male mating behavior / protein de-2-hydroxyisobutyrylase activity / protein lysine delactylase activity / negative regulation of dendritic spine development / p75NTR negatively regulates cell cycle via SC1 / fungiform papilla formation / epidermal cell differentiation / histone decrotonylase activity / NuRD complex / positive regulation of interleukin-1 production ...positive regulation of male mating behavior / protein de-2-hydroxyisobutyrylase activity / protein lysine delactylase activity / negative regulation of dendritic spine development / p75NTR negatively regulates cell cycle via SC1 / fungiform papilla formation / epidermal cell differentiation / histone decrotonylase activity / NuRD complex / positive regulation of interleukin-1 production / : / regulation of cell fate specification / EGR2 and SOX10-mediated initiation of Schwann cell myelination / negative regulation of stem cell population maintenance / regulation of stem cell differentiation / histone deacetylase activity, hydrolytic mechanism / histone deacetylase / ESC/E(Z) complex / cardiac muscle hypertrophy / cellular response to dopamine / positive regulation of intracellular estrogen receptor signaling pathway / behavioral response to ethanol / odontogenesis of dentin-containing tooth / STAT3 nuclear events downstream of ALK signaling / embryonic digit morphogenesis / histone deacetylase activity / protein lysine deacetylase activity / Hydrolases; Acting on carbon-nitrogen bonds, other than peptide bonds; In linear amides / response to caffeine / eyelid development in camera-type eye / Notch-HLH transcription pathway / positive regulation of oligodendrocyte differentiation / Sin3-type complex / dendrite development / positive regulation of stem cell population maintenance / histone deacetylase complex / response to amyloid-beta / positive regulation of proteolysis / progesterone receptor signaling pathway / RNA Polymerase I Transcription Initiation / hair follicle placode formation / response to hyperoxia / cellular response to transforming growth factor beta stimulus / Regulation of MECP2 expression and activity / FOXO-mediated transcription of oxidative stress, metabolic and neuronal genes / positive regulation of epithelial to mesenchymal transition / NF-kappaB binding / Transcriptional regulation of brown and beige adipocyte differentiation by EBF2 / cellular response to retinoic acid / Regulation of TP53 Activity through Acetylation / MECP2 regulates neuronal receptors and channels / heat shock protein binding / response to amphetamine / negative regulation of cell migration / SUMOylation of chromatin organization proteins / Regulation of PTEN gene transcription / transcription coregulator binding / ERCC6 (CSB) and EHMT2 (G9a) positively regulate rRNA expression / Regulation of endogenous retroelements by KRAB-ZFP proteins / response to nicotine / response to cocaine / negative regulation of neuron projection development / circadian regulation of gene expression / HDACs deacetylate histones / negative regulation of transforming growth factor beta receptor signaling pathway / Regulation of endogenous retroelements by Piwi-interacting RNAs (piRNAs) / promoter-specific chromatin binding / Negative Regulation of CDH1 Gene Transcription / NoRC negatively regulates rRNA expression / NOTCH1 Intracellular Domain Regulates Transcription / cellular response to hydrogen peroxide / Constitutive Signaling by NOTCH1 PEST Domain Mutants / Constitutive Signaling by NOTCH1 HD+PEST Domain Mutants / histone deacetylase binding / positive regulation of tumor necrosis factor production / cellular response to heat / Factors involved in megakaryocyte development and platelet production / response to lipopolysaccharide / heterochromatin formation / histone binding / Potential therapeutics for SARS / RNA polymerase II-specific DNA-binding transcription factor binding / proteasome-mediated ubiquitin-dependent protein catabolic process / chromosome, telomeric region / response to xenobiotic stimulus / chromatin remodeling / negative regulation of DNA-templated transcription / positive regulation of cell population proliferation / chromatin binding / negative regulation of apoptotic process / positive regulation of DNA-templated transcription / chromatin / negative regulation of transcription by RNA polymerase II / enzyme binding / positive regulation of transcription by RNA polymerase II / protein-containing complex / RNA binding / nucleoplasm / nucleus / cytoplasm
Similarity search - Function
Kelch repeat and BTB domain-containing protein 4 / KBTBD4, BTB/POZ domain / KBTBD4, BACK domain / Kelch repeat type 2 / Kelch motif / Histone deacetylase / BTB-kelch protein / BTB/Kelch-associated / BTB And C-terminal Kelch / BTB And C-terminal Kelch ...Kelch repeat and BTB domain-containing protein 4 / KBTBD4, BTB/POZ domain / KBTBD4, BACK domain / Kelch repeat type 2 / Kelch motif / Histone deacetylase / BTB-kelch protein / BTB/Kelch-associated / BTB And C-terminal Kelch / BTB And C-terminal Kelch / Histone deacetylase family / Kelch-type beta propeller / Histone deacetylase domain / Histone deacetylase domain superfamily / Histone deacetylase domain / Ureohydrolase domain superfamily / BTB/POZ domain / BTB domain profile. / Broad-Complex, Tramtrack and Bric a brac / BTB/POZ domain / SKP1/BTB/POZ domain superfamily
Similarity search - Domain/homology
Histone deacetylase 2 / Kelch repeat and BTB domain-containing protein 4
Similarity search - Component
Biological speciesHomo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.71 Å
AuthorsChen, Z. / Chi, G. / Pike, A.C.W. / Montes, B. / Bullock, A.N.
Funding support United Kingdom, Switzerland, 2items
OrganizationGrant numberCountry
Cancer Research UKDRCNPG-May21/100002 United Kingdom
Innovative Medicines Initiative875510 Switzerland
CitationJournal: Nat Commun / Year: 2025
Title: Structural mimicry of UM171 and neomorphic cancer mutants co-opts E3 ligase KBTBD4 for HDAC1/2 recruitment.
Authors: Zhuoyao Chen / Gamma Chi / Timea Balo / Xiangrong Chen / Beatriz Ralsi Montes / Steven C Clifford / Vincenzo D'Angiolella / Timea Szabo / Arpad Kiss / Tibor Novak / András Herner / András ...Authors: Zhuoyao Chen / Gamma Chi / Timea Balo / Xiangrong Chen / Beatriz Ralsi Montes / Steven C Clifford / Vincenzo D'Angiolella / Timea Szabo / Arpad Kiss / Tibor Novak / András Herner / András Kotschy / Alex N Bullock /
Abstract: Neomorphic mutations and drugs can elicit unanticipated effects that require mechanistic understanding to inform clinical practice. Recurrent indel mutations in the Kelch domain of the KBTBD4 E3 ...Neomorphic mutations and drugs can elicit unanticipated effects that require mechanistic understanding to inform clinical practice. Recurrent indel mutations in the Kelch domain of the KBTBD4 E3 ligase rewire epigenetic programs for stemness in medulloblastoma by recruiting LSD1-CoREST-HDAC1/2 complexes as neo-substrates for ubiquitination and degradation. UM171, an investigational drug for haematopoietic stem cell transplantation, was found to degrade LSD1-CoREST-HDAC1/2 complexes in a wild-type KBTBD4-dependent manner, suggesting a potential common mode of action. Here, we identify that these neomorphic interactions are mediated by the HDAC deacetylase domain. Cryo-EM studies of both wild-type and mutant KBTBD4 capture 2:1 and 2:2 KBTBD4-HDAC2 complexes, as well as a 2:1:1 KBTBD4-HDAC2-CoREST1 complex, at resolutions spanning 2.7 to 3.3 Å. The mutant and drug-induced complexes adopt similar structural assemblies requiring both Kelch domains in the KBTBD4 dimer for each HDAC2 interaction. UM171 is identified as a bona fide molecular glue binding across the ternary interface. Most strikingly, the indel mutation reshapes the same surface of KBTBD4 providing an example of a natural mimic of a molecular glue. Together, the structures provide mechanistic understanding of neomorphic KBTBD4, while structure-activity relationship (SAR) analysis of UM171 reveals analog S234984 as a more potent molecular glue for future studies.
History
DepositionAug 13, 2024Deposition site: PDBE / Processing site: PDBE
Revision 1.0Apr 9, 2025Provider: repository / Type: Initial release

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: Isoform 1 of Kelch repeat and BTB domain-containing protein 4
D: Histone deacetylase 2
C: Isoform 1 of Kelch repeat and BTB domain-containing protein 4
B: Histone deacetylase 2
hetero molecules


Theoretical massNumber of molelcules
Total (without water)227,9436
Polymers227,8124
Non-polymers1312
Water00
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: electron microscopy, not applicable
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1

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Components

#1: Protein Isoform 1 of Kelch repeat and BTB domain-containing protein 4 / BTB and kelch domain-containing protein 4


Mass: 58463.059 Da / Num. of mol.: 2 / Mutation: Indel mutation R313PRR
Source method: isolated from a genetically manipulated source
Details: Contains medulloblastoma cancer indel mutation R313PRR
Source: (gene. exp.) Homo sapiens (human) / Gene: KBTBD4, BKLHD4 / Cell line (production host): Expi293F / Production host: Homo sapiens (human) / References: UniProt: Q9NVX7
#2: Protein Histone deacetylase 2 / HD2 / Protein deacylase HDAC2


Mass: 55443.156 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: HDAC2 / Cell (production host): Expi293F / Production host: Homo sapiens (human)
References: UniProt: Q92769, histone deacetylase, Hydrolases; Acting on carbon-nitrogen bonds, other than peptide bonds; In linear amides
#3: Chemical ChemComp-ZN / ZINC ION


Mass: 65.409 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: Zn
Has ligand of interestN
Has protein modificationN

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

ComponentName: KBTBD4 R313PRR cancer mutant in complex with HDAC2 at 2:2 stoichiometry
Type: COMPLEX / Entity ID: #1-#2 / Source: RECOMBINANT
Molecular weightExperimental value: NO
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Organism: Homo sapiens (human)
Buffer solutionpH: 7.5
Buffer component
IDConc.NameFormulaBuffer-ID
150 mMHEPESC8H18N2O4S1
2150 mMsodium chlorideNaCl1
35 mMmagnesium dichlorideMgCl21
410 %glycerolC3H5(OH)31
50.5 mMTCEPC9H15O6P1
SpecimenConc.: 0.4 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: GOLD / Grid mesh size: 300 divisions/in. / Grid type: UltrAuFoil R1.2/1.3
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277.15 K

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal magnification: 105000 X / Nominal defocus max: 2800 nm / Nominal defocus min: 1000 nm
Specimen holderCryogen: NITROGEN
Image recordingElectron dose: 38.6 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Num. of grids imaged: 1 / Num. of real images: 13131
Image scansWidth: 5760 / Height: 4092

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Processing

EM softwareName: PHENIX / Version: 1.20.1_4487: / Category: model refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
SymmetryPoint symmetry: C2 (2 fold cyclic)
3D reconstructionResolution: 2.71 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 295217 / Symmetry type: POINT
Atomic model buildingB value: 107.7 / Protocol: FLEXIBLE FIT / Space: REAL
Atomic model building
IDPDB-ID 3D fitting-IDAccession codeInitial refinement model-IDSource nameType
17ZZT

7zzt

17ZZT1PDBexperimental model
21AF-Q9N010-F12AlphaFoldin silico model
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00411698
ELECTRON MICROSCOPYf_angle_d0.77415918
ELECTRON MICROSCOPYf_dihedral_angle_d3.9281672
ELECTRON MICROSCOPYf_chiral_restr0.0481828
ELECTRON MICROSCOPYf_plane_restr0.0062034

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