Discovering protein-protein interaction stabilisers by native mass spectrometry

Area of Science:

• Biochemistry
• Chemical Biology
• Structural Biology

Background:

• Protein-protein interactions (PPIs) are crucial therapeutic targets, with stabilization offering an alternative strategy to inhibition for disrupted PPIs in disease.
• Discovering PPI stabilizers is challenging due to the lack of tools to monitor stabilization, stoichiometry, and binding equilibria simultaneously.

Purpose of the Study:

• To demonstrate the utility of native mass spectrometry (MS) as a rapid screening tool for identifying protein-protein interaction (PPI) stabilizers.
• To simultaneously monitor binding stoichiometry and equilibria shifts induced by potential stabilizers.

Main Methods:

• Utilized native mass spectrometry (MS) to analyze three specific PPIs involving 14-3-3σ and its partners (ERα, p53, LRRK2).
• Assessed the differential stabilization effects of fusicoccin A on these PPIs within a single MS measurement.

Main Results:

• Native MS successfully determined the stoichiometry and binding equilibria for 14-3-3σ interactions with ERα, p53, and LRRK2.
• Observed a significant shift in binding equilibria for the 14-3-3σ:ERα complex upon fusicoccin A addition, distinct from the 14-3-3σ:p53 and 14-3-3σ:LRRK2 complexes.
• Gained insights into the dynamics of ternary complex formation and the differential modulation of PPIs by stabilizers.

Conclusions:

• Native MS is a powerful technique for rapidly screening and identifying PPI stabilizers.
• This method provides crucial information on binding stoichiometry, equilibria, and complex dynamics, highlighting its potential as a primary screening technology for novel therapeutics.