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Figure 4.
Figure 4. SPOP^BTB+ Forms a 2:2 Dimer with Cul3 N-Terminal
Domain
(A) Left, overall view of the SPOP^BTB+ dimer, with
protomers in cyan (A) and red (B). Right, close-up view of dimer
interface rotated 90° in x.
(B) Equilibrium AUC of
SPOP^BTB+ + Cul3^NTD. Samples at 1.0 to 8.8 μM centrifuged at
8,000 (red), 12,000 (blue), and 16,000 (black) rpm (4°C).
Lines show global nonlinear least-squares best-fit of all data
sets/concentrations/speeds to a heterogeneous association model
describing a 2:2 SPOP^BTB+:Cul3^NTD complex (MW 127.1 kDa) with
indicated K[D] value. For clarity, only the 3 μM sample is
shown.
(C) AUC of L186D, L190D, L193D, and I217K mutant
SPOP^BTB+ + Cul3^NTD performed as in (B). Lines show global
nonlinear least-squares best-fit of all data
sets/concentrations/ speeds to a heterogeneous association model
describing a 1:1 SPOP^BTB+ (mutant):Cul3^NTD complex (MW 63.6
kDa) with the indicated K[D] value. For clarity, only the 2.0
μM sample is shown.
(D) Western blots of
SPOP^MATH-BTB+-mediated ubiquitination detecting His-Puc for
wild-type and L186D, L190D, L193D, and I217K (dimer-defective)
mutant SPOP.
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