Deciphering Mode of Action of Functionally Important Regions in the Intrinsically Disordered Paxillin (Residues 1-313) Using Its Interaction with FAT (Focal Adhesion Targeting Domain of Focal Adhesion Kinase)
Fig 1
Dissection of paxillin constructs (residues 1–313) followed by expression and interaction studies.
(a) Timeline for overall-strategy. (b) Illustration of solubility and activity level of linear dissected human paxillin (residues 1–313). (c) Phosphor screen image of filter assay for optimization of temperature for paxillin constructs (left). Tabular representation of paxillin constructs, negative and positive controls corresponding to each well in filter assay [1]. (d) Phosphor screen image of 10% SDS PAGE of 35S labeled cell-free expressed samples after GST pull-down assay of the paxillin constructs A1–E1; The right panel shows fraction of interaction of each construct with respect to B2 (since B2 showed maximum level of interaction) (e) Illustration of solubility and activity of dissected C3 constructs. All experiments were performed in triplicates and averaged. To rule out non-specific interactions that might occur with GST tagged FAT, GFP that was expressed in cell-free system and a reaction without DNA template were used as negative controls.