Experiment-guided molecular simulations define a heterogeneous structural ensemble for the

SHP2 plays an important role in regulating cellular processes, and its pathogenic mutations cause developmental disorders and are linked to cancer. SHP2 is a multidomain protein, comprising two SH2 domains arranged in tandem, a catalytic PTP domain, and a disordered C-terminal tail. SHP2 is activated upon binding two linked phosphopeptides to its SH2 domains, and the peptide orientation and spacing between binding sites are critical for enzymatic activation. For decades, the tandem SH2 has been extensively studied to identify the relative orientation of the two SH2 domains that most effectively binds effectors. So far, neither crystallography nor experiments in solution have provided conclusive results. Using experiment-guided molecular simulations, we determine the heterogeneous structural ensemble of the tandem SH2 in solution in agreement with experimental data from small-angle X-ray scattering and NMR residual dipolar couplings. In the solution ensemble, N-SH2 adopts different orientations and positions relative to C-SH2. We suggest that the intrinsic structural plasticity of the tandem SH2 allows SHP2 to respond to external stimuli and is essential for its functional activity.