Reverse Binding Mode of Phosphotyrosine Peptides with SH2 Protein.

Discerning the different interaction states during dynamic protein-ligand binding is difficult. Here we apply site-specific cysteine-α-chloroacetyl cross-linking to scrutinize the binding between the Src homology 2 (SH2) domain and phosphotyrosine (pY) peptides, a highly dynamic interaction that is a key to cellular signal transduction. From a model SH2 protein to a set of representative SH2 domains, we showed here that a proximity-induced cysteine-α-chloroacetyl reaction cross-linked two spatially adjacent chemical groups as a result of the binding interaction, and reciprocally, the information about the interaction states can be deduced from the cross-linked products. To our surprise, we found SH2 domains can adopt a reverse binding mode with "single-pronged", "two-pronged", and "half" pY peptides. This finding was further supported by a set of 500 ns molecular dynamics simulations. This serendipitous finding defies the canonical theory of SH2 binding, suggests a possible answer about the source of the versatility of SH2 signaling, and sets a model for other protein binding interactions.

Switching substitution groups on the in-tether chiral centre influences backbone peptides' permeability and target binding affinity.

Different substitution groups on the in-tether chiral centre of chirality-induced helical peptides (CIH peptides) showed distinguishable effects on the peptides' cellular uptakes and binding affinities with the estrogen receptor α(ER-α). This study proves that in-tether chiral centres are a valuable modification site for constructing peptide ligands with preferable biophysical properties.