Hotspot Interactions between Two Fab Molecules in Molecular Dynamics Simulations Improve Predictive Models of...

Area of Science:

• Biochemistry and Molecular Biology
• Computational Biology
• Protein Engineering

Background:

• Protein-protein interactions (PPIs) are central to biological functions.
• Identifying interaction hotspots is key for understanding protein aggregation and engineering proteins.
• Limited research has explored the roles of surface vs. buried interactions (e.g., aggregation-prone regions or APRs).

Purpose of the Study:

• To investigate and characterize key interaction sites between two Fab antibody fragments using molecular dynamics (MD) simulations.
• To differentiate persistent interaction hotspots from transient contacts.
• To assess the impact of identified hotspots and APRs on predicting protein aggregation kinetics.

Main Methods:

• All-atom molecular dynamics (MD) simulations of two Fab antibody fragments.
• Frequency contact map analysis to identify stable residue contacts.
• Principal component analysis (PCA) to analyze conformational dynamics.
• Inclusion of solvent accessibility data for surface hotspots and predicted APRs.

Main Results:

• Few persistent interaction hotspots were identified, characterized by sustained stable contacts.
• Interactions between Fab fragments significantly influenced conformational dynamics compared to single fragment simulations.
• Incorporating solvent accessibility for surface hotspots and a predicted APR improved aggregation kinetics models across 49 conditions.

Conclusions:

• Persistent interaction sites, including buried APRs, play a critical role in protein aggregation.
• Molecular-level insights from MD simulations are valuable for protein engineering strategies.
• Modulating these identified interactions can enhance protein product stability and therapeutic efficacy.