Native MS and ligand observed NMR uncovers subtle SLiM binding variations that mediate HSP90-Hop PPI modulation

Area of Science:

• Biochemistry
• Structural Biology
• Drug Discovery

Background:

• Heat Shock Protein 90 (HSP90) is crucial for cellular proteostasis.
• HSP90 interacts with co-chaperones like Hop via its C-terminal MEEVD motif.
• Targeting HSP90-co-chaperone interactions is a promising therapeutic strategy.

Purpose of the Study:

• Investigate why a non-natural tetrazole peptide (2) inhibits HSP90-Hop interaction more effectively than the native MEEVD motif (1).
• Determine the structural basis for differential inhibition of protein-protein interactions (PPIs).

Main Methods:

• Native mass spectrometry (nMS) with ion mobility (IM).
• Saturation transfer difference (STD) NMR.
• Water ligand observed via gradient spectroscopy (WLOGSY) NMR.
• Analysis of peptide derivatives binding to the Hop_TPR2A domain.

Main Results:

• Peptide 2, mimicking the MEEVD motif, disrupts HSP90-Hop PPIs.
• Native MEEVD peptide (1) showed weak inhibition despite similar binding affinity.
• Differences in PPI modulation stem from altered binding modes, not affinity, due to sequence variations.
• Conformational stability and magnetization transfer varied subtly between peptides.

Conclusions:

• The study provides a structural basis for designing peptidomimetic inhibitors of HSP90-Hop PPIs.
• Altered binding modes, rather than affinity, are key to modulating transient PPIs.
• A generalized strategy for targeting transient PPIs in drug discovery is presented.