Extended Data Table 2 |.

Cryo-EM data collection, refinement, and validation statistics.

IκBα–UB~UBE2D crosslink	UBE2D2	UBE2D3	UBE2D3	UBE2D2	UBE2D2	none	none
(C21I C107A Cl 1 ID)	2×2way XL	3way XL	3way XL	3way XL	3way XL
Substrate Receptor	β-TRCP2	β-TRCP1	β-TRCP1	β-TRCP1	β-TRCP1	β-TRCP1	β-TRCP1
		175-C	175-C	175-C	175-C	175-C	175-C
RBX1	WT	WT	N98R	N98R	N98R	WT	WT
NEDD8	yes	yes	yes	yes	yes	no	yes
SKP1	WT	ΔΔ	ΔΔ	ΔΔ	ΔΔ	WT	WT
	EMDB-10578	EMDB-10579	EMDB-10580	EMDB-10581	EMDB-10585	EMDB-10582	EMDB-10583
					PDB 6TTU
Data collection and processing
Microscope	Krios	Arctica	Arctica	Arctica	Krios	Glacios	Glacios
Magnification	105,000	92,000	73,000	73,000	130,000	36,000	22,000
Voltage (kV)	300	200	200	200	300	200	200
Electron exposure (e−/Å2)	56	61.3	60.8	70	70.2	60	59
Defocus range (μm)	−1.2 ~ −3.6	−1.5 ~ −3.5	−1.5 ~ −3.5	−1.5 ~ −3.5	−1.2 ~ −3.3	−1.2 ~ −3.3	−1.2 ~ −3.3
Pixel size (Å)	1.34	1.612	1.997	1.997	1.06	1.181	1.885
Symmetiy imposed	C2	Cl	Cl	Cl	Cl	Cl	Cl
Initial particle images (no.)	2,575,161	464,344	601,121	459,011	1,661,870	2,051,804	1,666,293
Final particle images (no.)	33,738	47,246	107,311	40,835	106,257	262,116	349,803
Map resolution (Å)	9.3	8.6	9.4	8.4	3.72	4.64	6.7
FSC threshold	(0.143)	(0.143)	(0.143)	(0.143)	(0.143)	(0.143)	(0.143)
Map resolution range (Å)	-	-	-	-	3.46~6.0	-	-
Refinement
Initial model used					1LDJ 1P22
(PDB code)					4P5O 4V3L
Model resolution (Å)					3.7
FSC threshold					(0.143)
Map sharpening B factor (Å2)	−578.9	−1159	−1272	−983.5	−94.2	−199	−338
Model composition
Non-hydrogen atoms					13034
Protein residues					1616
Ligands					3(ZN)
B factors (Å2)
Protein					91
Ligand					224
R.m.s. deviations
Bond lengths (Å)					0.011
Bond angles (°)					1.043
Validation
MolProbity score					2.37
Clashscore					16.31
Poor rotamers (%)					0.21
Ramachandran plot
Favored (%)					85.12
Allowed (%)					14.88
Disallowed (%)					0