Targeting disordered-structured domain interactions in Galectin-3 based on NMR and enhanced MD

Area of Science:

• Biochemistry and Structural Biology
• Drug Discovery and Development
• Computational Biology

Background:

• Intrinsically disordered regions (IDRs) are crucial functional protein domains but challenging for drug development due to their lack of defined structures.
• Galectin-3, implicated in cancer and inflammation, possesses a functionally important disordered N-terminal domain (NTD) that interacts with its C-terminal domain (CTD).
• Targeting the dynamic NTD of Galectin-3 for therapeutic intervention is hindered by structural ambiguity.

Purpose of the Study:

• To overcome the challenges of targeting intrinsically disordered regions (IDRs) for drug development.
• To identify a druggable pocket within the C-terminal domain (CTD) of Galectin-3 that interacts with its intrinsically disordered N-terminal domain (NTD).
• To design and validate peptide inhibitors targeting the newly discovered binding site in Galectin-3.

Main Methods:

• Integration of nuclear magnetic resonance (NMR) data from recombinant Galectin-3 with accelerated molecular dynamics (MD) simulations.
• In silico screening of candidate inhibitory peptides targeting the identified CTD pocket.
• Experimental validation of peptide inhibitors using agglutination assays and NMR spectroscopy.

Main Results:

• A specific pocket in the CTD of Galectin-3 was identified through combined NMR and MD simulations, showing frequent contact with the NTD.
• Mutational analysis confirmed the functional importance of this pocket by demonstrating loss of Galectin-3 agglutination activity.
• One designed peptide inhibitor effectively blocked Galectin-3 agglutination by binding to the CTD cavity, not the carbohydrate-recognition/binding domain (CRD).

Conclusions:

• A synergistic approach using MD simulations and NMR experiments can accurately predict binding interfaces between disordered and structured protein domains.
• This methodology enables the rational design of inhibitors for challenging targets involving intrinsically disordered regions (IDRs).
• The developed strategy holds potential for broader application in developing therapeutics for various diseases linked to IDR interactions.