Fig 5. ‘KDSAF’ peptides bind to the ACR4 kinase.

(A) Schematics of the ACR4, CRK1 and CRR3 amino acid sequence. The location of the “KDSAF” motif sequence in the domain architecture of the respective proteins is indicated by an arrow. (B) The ‘KDSAF’ motif containing peptides were incubated with the sACR4 KC protein to demonstrate specificity of binding towards the ACR4 kinase. A peptide overlay assay showing that the unphosphorylated JMD1 [26], the CRR3 hC and CRK1 hC peptides can bind to sACR4 KC (upper panel). Control blots show no specificity towards the SUMO solubility tag (lower panel). (C) Peptide binding assay demonstrating the ‘KDSAF’ containing peptides can bind to the natively folded sACR4 KC protein immobilized on a Ni²⁺ coated plate. Each peptide binding experiment was performed in triplicate and data represented as the mean ± the S.E. Data were corrected for dilution factor and subtracting the absorbance of the control (SUMO) from the experimental samples.