Supplementary Figure 5: Characterization of the conformational states of cIAP1 variants.

(a) Size-exclusion multi-angle light scattering (SEC-MALS) traces of cIAP1 constructs with and without SMAC mimetics. Apo-proteins are shown in black, AVPW-bound protein in blue, and BV6 bound proteins (as a dimerization control) in red. The UV absorbance for each peak was normalized to 1. As expected, cIAP1-B3R (top) dimerizes in response to both AVPW and BV6, while cIAP1-B3R-ΔCARD (center) is constitutively open and dimerizes only in the presence of BV6. The L617E construct (bottom) blocks AVPW-induced dimerization without disrupting the closed conformation. (b) L617 is positioned in the RING dimerization interface. The structure of the cIAP2 RING dimer (PDB code: 3EB5, ref. 4) is shown in cartoon representation with the homologous residue to L617, L603, colored magenta and shown as spheres. Zinc ions are shown as light gray spheres.