Mutation-induced rigidity in the Fyn SH2 domain enhances pY-binding affinity at the cost of peptide specificity

Abstract

Interactions between SH2 domains and tyrosine-phosphorylated (pY) peptides are essential for cellular signaling. While structural studies have revealed how triple-point Fyn SH2 mutants achieve ultra-high pY-peptide affinity, the dynamic consequences of these mutations remain unexplored. In this study, we performed extensive all-atom molecular dynamics simulations on the isolated wild-type Fyn SH2 domain, its mutant, and their complexes with the pY-peptide (EPQpYEEIPIYL). Comparative analyses of these simulations provided dynamic insights into how mutations within the pY-binding pocket alter the interaction between Fyn SH2 domain and the pY-peptide. Our results demonstrate that the mutations significantly influence the dynamic stability of unstructured regions within the SH2 domain and the domain-peptide interface. Specifically, the mutations enhance the rigidity and stability of the pY-binding pocket, as well as the overall structural stability of the domain, including the central β-sheet and terminal regions. This increased rigidity in the mutant enhances interactions between the pY-binding pocket and pY but weakens the interaction with the peptide residue at the +3 position relative to pY, thereby compromising the specificity of the domain-peptide interaction. These findings highlight that the interaction between SH2 domains and pY-peptides is governed not only by the structural properties of the pY-binding pocket but also by the dynamic stability of the domain itself. This insight could guide the experimental design of SH2 domains engineered to recognize post-translational modifications with diverse characteristics.