SH2 domains are structural modules specialized in the recognition and binding of target sequences containing a phosphorylated tyrosine residue. They are mostly incorporated in the 3D structure of scaffolding proteins that represent fundamental regulators of several signaling pathways. Among those, Crkl plays key roles in cell physiology by mediating signals from a wide range of stimuli, and its overexpression is associated with several types of cancers. In myeloid cells expressing the oncogene BCR/ABL, one interactor of Crkl-SH2 is the focal adhesion protein Paxillin, and this interaction is crucial in leukemic transformation. In this work, we analyze both the folding pathway of Crkl-SH2 and its binding reaction with a peptide mimicking Paxillin, under different ionic strength and pH conditions, by using means of fluorescence spectroscopy. From a folding perspective, we demonstrate the presence of an intermediate along the reaction. Moreover, we underline the importance of the electrostatic interactions in the early event of recognition, occurring between the phosphorylated tyrosine of the Paxillin peptide and the charge residues of Crkl-SH2. Finally, we highlight a pivotal role of a highly conserved histidine residue in the stabilization of the binding complex. The experimental results are discussed in light of previous works on other SH2 domains.
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