Native dynamics and allosteric responses in PTP1B probed by high-resolution HDX-MS.

Area of Science:

• Biophysics
• Structural Biology
• Pharmacology

Background:

• Protein tyrosine phosphatase 1B (PTP1B) is a key therapeutic target for metabolic diseases and cancer.
• Understanding PTP1B's conformational dynamics and allostery is crucial for developing effective allosteric inhibitors.
• Current knowledge of PTP1B allostery is limited, hindering inhibitor development.

Purpose of the Study:

• To investigate the solution dynamics and allosteric responses of PTP1B using high-resolution hydrogen-deuterium exchange mass spectrometry (HDX-MS).
• To map the effects of active-site and allosteric small-molecule inhibitors on PTP1B dynamics.
• To elucidate the mechanisms of allosteric inhibition in PTP1B.

Main Methods:

• High-resolution hydrogen-deuterium exchange mass spectrometry (HDX-MS) was employed to map backbone amide exchange rates.
• HDX-MS data was used to assess solution dynamics of apo PTP1B and PTP1B bound to inhibitors.
• HDX-MS results were compared with data from crystal structures and other biophysical techniques.

Main Results:

• HDX-MS revealed flexible loops and rigid regions in apo PTP1B, providing insights into its conformational heterogeneity.
• Inhibitor binding, particularly allosteric inhibitors, induced widespread changes in PTP1B dynamics, extending beyond known allosteric networks.
• Allosteric inhibitors affected dynamics in regions distal (>35 Å) from the ligand binding sites, suggesting a novel allosteric mechanism.

Conclusions:

• HDX-MS is a powerful tool for studying protein dynamics and allosteric effects of small-molecule ligands.
• Allosteric inhibition of PTP1B involves complex conformational changes, including distal residue modulation for entropic balancing.
• Integrated approaches combining HDX-MS with crystallography, NMR, and simulations can guide the development of novel PTP1B therapeutics.