Fig. 4: PEAK3:14-3-3 forms a stable high affinity heterocomplex. | Nature Communications

Fig. 4: PEAK3:14-3-3 forms a stable high affinity heterocomplex.

From: Structural mapping of PEAK pseudokinase interactions identifies 14-3-3 as a molecular switch for PEAK3 signaling

Fig. 4

a Recombinant PEAK3FL from insect cells elutes on size exclusion chromatography (SEC; S200 10/300) as a high affinity stoichiometric complex of dimeric PEAK3 with a 14-3-3ε,ζ heterodimer, results supported by SDS-PAGE analysis of eluted fractions (n = 3, independent experiments) (see also Supplementary Fig. 4a, source data). b Immunoprecipitation of PEAK3 WT and PEAK3 S69A showing S69 is required for 14-3-3 co-immunoprecipitation from cells (n = 3 biologically independent samples, see Supplementary Fig. 4a for biological repeats). c SEC-MALS analysis of PEAK3FL−14-3-3ε,ζ heterodimer complex confirming the experimentally determined mass closely matches the mass of 155 kDa expected for a stoichiometric dimer:dimer complex (n = 3, independent experiments). d SEC profile (S200 10/300) of recombinant PEAK3FL−14-3-3ε,ζ heterodimer pre-incubated with CrkIIFL dimer in a 1:1.2 molar ratio showing no complex formation (n = 3, independent experiments). Source data are provided as a source data file.

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