Figure 5: Molecular modelling and mutagenesis identify the predominant orientation for the procaspase 8–FADD interaction. | Nature Communications

Figure 5: Molecular modelling and mutagenesis identify the predominant orientation for the procaspase 8–FADD interaction.

From: Differential affinity of FLIP and procaspase 8 for FADD’s DED binding surfaces regulates DISC assembly

Figure 5

(a) Homology model of human procaspase 8 DED1 and DED2 (light and dark magenta) superimposed on vFLIP X-ray protein (PDB id 2BBR; light green and green) and α3 region of FADD NMR-solved structure (PDB id 2GF5; dark green). The procaspase 8 DED1/2-MC159 identity is 29%; the r.m.s.d. calculated on Cα atoms is 1.03 Å. Two possible orientations of the procaspase 8/FADD interaction and their predicted docking energies are depicted: (b) F122 located on α2 helix of procaspase 8 DED2 is accommodated into a hydrophobic groove formed between α1 and α4 helices of FADD DED; (c) F25 located on α2 helix of FADD DED is accommodated into a hydrophobic groove formed between α1 and α4 helices of procaspase 8 DED1. (d) Western blot analysis of GST pull-down assay using FADD DED as bait and wild-type and mutant Flag-procaspase 8 constructs as prey. (e) Western blot analysis of a GST pull-down competition experiment, in which binding of Flag-tagged wild-type procaspase 8 and Flag-tagged wild-type FLIP(S) to the FADD DED was assessed in the presence or absence of the other protein. Procaspase 8 and FLIP(S) inputs were equalized prior to incubation with GST-FADD, and analysis of the first washes indicates that the pull downs were carried out with an excess of both Flag-tagged proteins.

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