# Phosphatase SHP2 pathogenic mutations enhance activity by altering conformational sampling

## TL;DR

This study explains how specific mutations in the SHP2 protein cause it to become overactive by changing its shape, leading to stronger binding of signaling molecules.

## Contribution

The study reveals a novel mechanism by which SHP2 mutations enhance activity through altered conformational dynamics.

## Key findings

- The T42A mutation increases phosphopeptide binding affinity by stabilizing a zipped β-sheet conformation.
- Conformational changes in SHP2's N-SH2 domain are essential for phosphopeptide recognition and regulation.
- Complementary structural and dynamic methods uncover how SH2 domains regulate signaling.

## Abstract

Here, we characterize how two pathogenic SH2-domain mutations alter SHP2 regulation and lead to hyperactivation. We identify an apo conformation of the N-SH2 domain in which Tyr66 occludes the peptide-binding cleft, indicating that a conformational change is required for full binding of activating phosphopeptides. Our data suggest that the T42A mutation shifts the equilibrium toward a zipped central β-sheet state in the peptide-bound N-SH2 domain as the most likely model underlying the measured 10-fold increased binding affinity. These results help clarify the structural basis for SHP2 regulation and illustrate how conformational dynamics shape SH2-phosphopeptide recognition.

SH2 domains are critical mediators of cellular signaling, although the molecular mechanisms by which they bind their phosphopeptide ligands remain incompletely understood. We investigate the atomic mechanisms underlying both healthy regulation and dysregulation of the human protein tyrosine phosphatase SHP2, a key regulator of cellular signaling. While most pathogenic mutations cluster near the PTP/N-SH2 interface, the E139D and T42A mutations are located within the regulatory SH2 domains, and their mechanisms of dysregulation remain controversial. The T42A mutation in the N-SH2 domain paradoxically increases phosphotyrosine-peptide binding affinity despite disrupting the hydrogen bond of T42 to the phosphoryl group, a puzzling contradiction that remains unresolved. We find that the T42A mutation shifts the conformational ensemble of peptide-bound N-SH2 toward a zipped β-sheet state and suppresses millisecond conformational exchange, supporting a model in which enhanced stabilization of the zipped conformation contributes to hyperactivation. This conformational shift provides a structural rationale for the increased affinity of T42A and helps reconcile previously conflicting models of peptide-induced SHP2 activation. By integrating X-ray ensemble refinement with NMR relaxation, our work illustrates how complementary structural and dynamic approaches can uncover regulatory mechanisms in SHP2 and may inform broader principles of SH2-mediated phosphopeptide recognition.