PDB-29hg: Cryo-EM structure of the CUL1-RBX1-SKP1-FBXO22 SCF ubiquition lig...

Basic information

• Entry: Database: PDB / ID: 29hg
• Title: Cryo-EM structure of the CUL1-RBX1-SKP1-FBXO22 SCF ubiquition ligase in complex with NSD2 via UNC10088

Components

• Cullin-1
• E3 ubiquitin-protein ligase RBX1
• F-box only protein 22
• Histone-lysine N-methyltransferase NSD2
• S-phase kinase-associated protein 1

• Keywords: LIGASE / Ubiquitin Ligase / Targeted protein degradation / cullin-RING ligase / SKP1 CUL1 F box SCF / Nuclear receptor binding SET domain-containing 2 NSD2

Function / homology

Function:

atrial septum secundum morphogenesis / [histone H3]-lysine36 N-dimethyltransferase / regulation of skeletal muscle fiber development / histone H4K20 methyltransferase activity / regulation of double-strand break repair via nonhomologous end joining / histone H3K36 dimethyltransferase activity / histone H3K36 trimethyltransferase activity / Parkin-FBXW7-Cul1 ubiquitin ligase complex / positive regulation of isotype switching to IgA isotypes / atrial septum primum morphogenesis / membranous septum morphogenesis / F-box domain binding / regulation of establishment of protein localization / PcG protein complex / histone H3K36 methyltransferase activity / negative regulation of beige fat cell differentiation / cullin-RING-type E3 NEDD8 transferase / NEDD8 transferase activity / cullin-RING ubiquitin ligase complex / maintenance of protein location in nucleus / Cul7-RING ubiquitin ligase complex / cellular response to chemical stress / Loss of Function of FBXW7 in Cancer and NOTCH1 Signaling / positive regulation of protein autoubiquitination / RNA polymerase II transcription initiation surveillance / protein neddylation / ubiquitin ligase activator activity / NEDD8 ligase activity / protein K27-linked ubiquitination / negative regulation of response to oxidative stress / nucleocytoplasmic transport / histone H3 methyltransferase activity / VCB complex / Cul5-RING ubiquitin ligase complex / ubiquitin-ubiquitin ligase activity / ubiquitin-dependent protein catabolic process via the C-end degron rule pathway / SCF ubiquitin ligase complex / Cul2-RING ubiquitin ligase complex / Cul3-RING ubiquitin ligase complex / negative regulation of type I interferon production / SCF-dependent proteasomal ubiquitin-dependent protein catabolic process / Cul4A-RING E3 ubiquitin ligase complex / Cul4-RING E3 ubiquitin ligase complex / Prolactin receptor signaling / negative regulation of mitophagy / Cul4B-RING E3 ubiquitin ligase complex / ubiquitin ligase complex scaffold activity / cullin family protein binding / protein monoubiquitination / ubiquitin-like ligase-substrate adaptor activity / site of DNA damage / signal transduction in response to DNA damage / Nuclear events stimulated by ALK signaling in cancer / protein K48-linked ubiquitination / transcription-coupled nucleotide-excision repair / negative regulation of insulin receptor signaling pathway / regulation of cellular response to insulin stimulus / positive regulation of TORC1 signaling / post-translational protein modification / intrinsic apoptotic signaling pathway / molecular function activator activity / animal organ morphogenesis / T cell activation / cellular response to starvation / protein modification process / Regulation of BACH1 activity / MAP3K8 (TPL2)-dependent MAPK1/3 activation / negative regulation of canonical NF-kappaB signal transduction / SCF-beta-TrCP mediated degradation of Emi1 / NIK-->noncanonical NF-kB signaling / Nonhomologous End-Joining (NHEJ) / cellular response to amino acid stimulus / Vpu mediated degradation of CD4 / Dectin-1 mediated noncanonical NF-kB signaling / Degradation of DVL / Degradation of CRY and PER proteins / G1/S transition of mitotic cell cycle / negative regulation of canonical Wnt signaling pathway / Activation of NF-kappaB in B cells / Iron uptake and transport / Degradation of GLI1 by the proteasome / GSK3B and BTRC:CUL1-mediated-degradation of NFE2L2 / Recognition of DNA damage by PCNA-containing replication complex / Negative regulation of NOTCH4 signaling / RING-type E3 ubiquitin transferase / G2/M DNA damage checkpoint / Hedgehog 'on' state / Vif-mediated degradation of APOBEC3G / FBXL7 down-regulates AURKA during mitotic entry and in early mitosis / bone development / Ubiquitin-Mediated Degradation of Phosphorylated Cdc25A / Degradation of GLI2 by the proteasome / GLI3 is processed to GLI3R by the proteasome / PKMTs methylate histone lysines / beta-catenin binding / NOTCH1 Intracellular Domain Regulates Transcription / Degradation of beta-catenin by the destruction complex / Evasion by RSV of host interferon responses / DNA Damage Recognition in GG-NER / Oxygen-dependent proline hydroxylation of Hypoxia-inducible Factor Alpha

Domain/homology:

FIST, C-domain / FIST C domain / FIST_C / : / : / : / : / : / : / : / : / F-box domain / NSD, Cys-His rich domain / : / : / : / NSD Cys-His rich domain / Histone-lysine N-methyltransferase NSD-like, PHD zinc finger / Histone-lysine N-methyltransferase NSD-like, variant PHD zinc finger / Histone-lysine N-methyltransferase NSD-like, PHD zinc finger 1 / : / AWS domain / AWS domain / AWS domain profile. / associated with SET domains / F-box-like domain superfamily / SKP1 component, dimerisation / S-phase kinase-associated protein 1 / SKP1-like, dimerisation domain superfamily / Skp1 family, dimerisation domain / F-box domain / Zinc finger, RING-H2-type / RING-H2 zinc finger domain / Cysteine-rich motif following a subset of SET domains / Cullin, N-terminal / Cullin protein neddylation domain / : / Cullin, conserved site / Cullin alpha solenoid domain / Cullin family signature. / Cullin repeat-like-containing domain superfamily / Cullin protein, neddylation domain / Cullin / Cullin protein neddylation domain / Post-SET domain / Post-SET domain profile. / Cullin / HMG (high mobility group) box / : / Cullin alpha+beta domain / Cullin homology domain / Cullin homology domain superfamily / Cullin family profile. / HMG boxes A and B DNA-binding domains profile. / high mobility group / High mobility group box domain / High mobility group box domain superfamily / S-phase kinase-associated protein 1-like / SKP1 component, POZ domain / Skp1 family, tetramerisation domain / Found in Skp1 protein family / SET (Su(var)3-9, Enhancer-of-zeste, Trithorax) domain / SET domain / SET domain profile. / SET domain superfamily / SET domain / domain with conserved PWWP motif / PWWP domain / PWWP domain profile. / PWWP domain / Zinc finger, PHD-type, conserved site / PHD-finger / Zinc finger PHD-type signature. / SKP1/BTB/POZ domain superfamily / Ring finger / Zinc finger PHD-type profile. / Zinc finger, PHD-finger / Zinc finger, PHD-type / PHD zinc finger / Zinc finger, FYVE/PHD-type / Zinc finger RING-type profile. / Zinc finger, RING-type / Zinc finger, RING/FYVE/PHD-type / Winged helix DNA-binding domain superfamily / Winged helix-like DNA-binding domain superfamily

Component:

: / Histone-lysine N-methyltransferase NSD2 / E3 ubiquitin-protein ligase RBX1 / S-phase kinase-associated protein 1 / Cullin-1 / F-box only protein 22

• Biological species: Homo sapiens (human)
• Method: ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 4 Å
• Authors: Amann, S.J. / Robertson, K.C. / Grishkovskaya, I. / Liu, T. / James, L.I. / Brown, N.B. / Haselbach, D.

Funding support

United States, 2items

Organization	Grant number	Country
National Institutes of Health/National Cancer Institute (NIH/NCI)	R35GM128855	United States
National Institutes of Health/National Cancer Institute (NIH/NCI)	R01CA242305	United States

Citation

Journal: Nat Commun / Year: 2026
Title: Structural basis of NSD2 degradation via targeted recruitment of SCF-FBXO22.
Authors: Kevin C Robertson / Sascha J Amann / Tongkun Liu / Adam V Funk / Xianxi Wang / Irina Grishkovskaya / Aamir Mehmood / John R Tabor / Jacqueline L Norris-Drouin / Cheryl H Arrowsmith / Jon L Collins / Yinglong Miao / Michael J Emanuele / David Haselbach / Lindsey I James / Nicholas G Brown /
Abstract: Targeted protein degradation (TPD) through the ubiquitin-proteasome system is driven by compound-mediated polyubiquitination of a protein-of-interest by an E3 ubiquitin (Ub) ligase. Relatively few E3s have been successfully utilized for TPD and the governing principles of functional ternary complex formation between the E3, degrader, and protein target remain elusive. FBXO22 has recently been harnessed for TPD applications by degraders that covalently modify its cysteine residues. Here, we reveal that the aldehyde derivative of UNC10088 promotes cooperative binding of FBXO22 to NSD2, a histone methyltransferase and oncogenic protein, leading to a cryo-EM structure of the SKP1-CUL1-F-box (SCF)-FBXO22 complex with NSD2. This structure revealed a conformational change in the FBXO22 loop surrounding C326, further exposing the cysteine for covalent recruitment. Additional medicinal chemistry efforts led to the discovery of benzaldehyde-based non-prodrug degraders that similarly engage C326 of FBXO22 and potently degrade NSD2. Unlike many degraders, our molecules recruit NSD2 to a different surface of FBXO22 than the known FBXO22 substrate BACH1, allowing for concurrent complex formation and structural determination of SCF bound to both the neosubstrate NSD2 and native substrate BACH1. Overall, we demonstrate the biochemical and structural basis for NSD2 degradation, revealing key principles for efficient and selective TPD by SCF.

#1: Journal: Acta Crystallogr D Struct Biol / Year: 2019
Title: Macromolecular structure determination using X-rays, neutrons and electrons: recent developments in Phenix.
Authors: Dorothee Liebschner / Pavel V Afonine / Matthew L Baker / Gábor Bunkóczi / Vincent B Chen / Tristan I Croll / Bradley Hintze / Li Wei Hung / Swati Jain / Airlie J McCoy / Nigel W Moriarty / Robert D Oeffner / Billy K Poon / Michael G Prisant / Randy J Read / Jane S Richardson / David C Richardson / Massimo D Sammito / Oleg V Sobolev / Duncan H Stockwell / Thomas C Terwilliger / Alexandre G Urzhumtsev / Lizbeth L Videau / Christopher J Williams / Paul D Adams /
Abstract: Diffraction (X-ray, neutron and electron) and electron cryo-microscopy are powerful methods to determine three-dimensional macromolecular structures, which are required to understand biological processes and to develop new therapeutics against diseases. The overall structure-solution workflow is similar for these techniques, but nuances exist because the properties of the reduced experimental data are different. Software tools for structure determination should therefore be tailored for each method. Phenix is a comprehensive software package for macromolecular structure determination that handles data from any of these techniques. Tasks performed with Phenix include data-quality assessment, map improvement, model building, the validation/rebuilding/refinement cycle and deposition. Each tool caters to the type of experimental data. The design of Phenix emphasizes the automation of procedures, where possible, to minimize repetitive and time-consuming manual tasks, while default parameters are chosen to encourage best practice. A graphical user interface provides access to many command-line features of Phenix and streamlines the transition between programs, project tracking and re-running of previous tasks.

History

Deposition	Mar 11, 2026	Deposition site: PDBE / Processing site: PDBE
Revision 1.0	May 6, 2026	Provider: repository / Type: Initial release
Revision 1.0	May 6, 2026	Data content type: EM metadata / Data content type: EM metadata / Provider: repository / Type: Initial release
Revision 1.0	May 6, 2026	Data content type: Image / Data content type: Image / Provider: repository / Type: Initial release
Revision 1.0	May 6, 2026	Data content type: Primary map / Data content type: Primary map / Provider: repository / Type: Initial release

Assembly

Deposited unit

A: F-box only protein 22

B: Histone-lysine N-methyltransferase NSD2

C: S-phase kinase-associated protein 1

D: Cullin-1

E: E3 ubiquitin-protein ligase RBX1

hetero molecules

Summary:

• 318 kDa, 5 polymers

Component details:

	Theoretical mass	Number of molelcules
Total (without water)	318,457	6
Polymers	317,848	5
Non-polymers	609	1
Water	0	0

1

Summary:

• Idetical with deposited unit
• defined by author&software
• Evidence: electron microscopy, not applicable

Symmetry operations:

Type	Name	Symmetry operation	Number
identity operation	1_555	x,y,z	1

Components

Protein , 5 types, 5 molecules A B C D E

#1: Protein

A F-box only protein 22 / F-box protein FBX22p44

Details:

Mass: 44562.270 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: FBXO22, FBX22 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q8NEZ5

#2: Protein

B Histone-lysine N-methyltransferase NSD2 / Multiple myeloma SET domain-containing protein / MMSET / Nuclear SET domain-containing protein 2 / Protein trithorax-5 / Wolf-Hirschhorn syndrome candidate 1 protein

Details:

Mass: 152515.469 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: NSD2, KIAA1090, MMSET, TRX5, WHSC1 / Production host: Trichoplusia (butterflies/moths)
References: UniProt: O96028, [histone H3]-lysine36 N-dimethyltransferase

#3: Protein

C S-phase kinase-associated protein 1 / Cyclin-A/CDK2-associated protein p19 / p19A / Organ of Corti protein 2 / OCP-2 / Organ of Corti protein II / OCP-II / RNA polymerase II elongation factor-like protein / SIII / Transcription elongation factor B polypeptide 1-like / p19skp1

Details:

Mass: 18679.965 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: SKP1, EMC19, OCP2, SKP1A, TCEB1L / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: P63208

#4: Protein

D Cullin-1 / CUL-1

Details:

Mass: 89800.367 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: CUL1 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q13616

#5: Protein

E E3 ubiquitin-protein ligase RBX1 / E3 ubiquitin-protein transferase RBX1 / Protein ZYP / RING finger protein 75 / RING-box protein 1 / Rbx1 / Regulator of cullins 1 / ROC1

Details:

Mass: 12289.977 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: RBX1, RNF75, ROC1 / Production host: Trichoplusia ni (cabbage looper)
References: UniProt: P62877, RING-type E3 ubiquitin transferase, cullin-RING-type E3 NEDD8 transferase

Non-polymers , 1 types, 1 molecules

#6: Chemical

B-1401 ChemComp-A1J20 / ~{N}-cyclopropyl-~{N}-[[4-[[2-(6-oxidanylhexanoyl)-1~{H}-isoquinolin-6-yl]carbamoyl]phenyl]methyl]-3-oxidanylidene-4~{H}-1,4-benzoxazine-7-carboxamide

Details:

Mass: 608.684 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C₃₅H₃₆N₄O₆ / Feature type: SUBJECT OF INVESTIGATION

Details

• Has ligand of interest: Y
• Has protein modification: N

Experimental details

Experiment

• Experiment: Method: ELECTRON MICROSCOPY
• EM experiment: Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

Sample preparation

• Component: Name: SCF FBXO22, NSD2, UNC10088 / Type: COMPLEX / Entity ID: #1-#5 / Source: RECOMBINANT
• Molecular weight: Experimental value: NO
• Source (natural): Organism: Homo sapiens (human)
• Source (recombinant): Organism: Trichoplusia ni (cabbage looper)
• Buffer solution: pH: 8

Buffer component

ID	Conc.	Name	Formula	Buffer-ID
1	20 mM	HEPES		1
2	200 mM	Sodium Chloride	NaCl	1
3	1 mM	DTT		1

• Specimen: Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
• Specimen support: Grid material: COPPER / Grid mesh size: 200 divisions/in. / Grid type: Quantifoil R1.2/1.3
• Vitrification: Instrument: LEICA EM GP / Cryogen name: ETHANE / Humidity: 75 % / Chamber temperature: 277 K

Electron microscopy imaging

• Experimental equipment: Model: Titan Krios / Image courtesy: FEI Company
• Microscopy: Model: TFS KRIOS
• Electron gun: Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
• Electron lens: Mode: BRIGHT FIELD / Nominal defocus max: 2000 nm / Nominal defocus min: 1000 nm / Alignment procedure: COMA FREE
• Specimen holder: Cryogen: NITROGEN
• Image recording: Electron dose: 50 e/Ų / Film or detector model: FEI FALCON IV (4k x 4k)

Processing

EM software

ID	Name	Category
1	cryoSPARC	particle selection
12	cryoSPARC	3D reconstruction
13	PHENIX	model refinement

• CTF correction: Type: PHASE FLIPPING AND AMPLITUDE CORRECTION
• 3D reconstruction: Resolution: 4 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 207711 / Symmetry type: POINT
• Atomic model building: Protocol: FLEXIBLE FIT / Space: REAL
• Atomic model building: Source name: AlphaFold / Type: in silico model