Structural Analysis of Integrin β3 Binding to the SH3 Domain of Src Kinase

Abstract

Src kinase plays an important role in integrin signaling by regulating cytoskeletal organization and cell remodeling. Previous in vivo studies have revealed that the SH3 domain of c-Src kinase directly associates with the C-terminus of β3 integrin cytoplasmic tail. Here we explore this binding interface with a combination of different spectroscopic and computational methods. Our NMR chemical shifts mapping data in concert with PRE and transferred NOE studies have confirmed the RGT762 motif of β3 integrin as the binding site for Src kinase. Our circular dichroism data shows that the C-terminus of β3 adopts a partial polyproline type II helix, a characteristic feature of SH3 canonical binding ligands. We have used our experimental data/restraints to generate a reliable model of the complex through docking. This model suggests a different binding mode from the one proposed through previous studies, wherein the C-terminal end of β3 spans the region in between the RT and n-Src loops of SH3 domain. Furthermore, we show that tyrosine phosphorylation of β3 prevents this interaction, supporting the notion of a constitutive interaction between resting β3 integrin and Src kinase.