Exploring Trop-2-Nanobody PPI Interactions through Molecular Dynamics Simulations and Biovalidations.

Area of Science:

• Biochemistry
• Molecular Biology
• Computational Biology

Background:

• Trop-2 is a transmembrane glycoprotein implicated in various cancers, making it a target for cancer therapy.
• Trop-2-specific antibodies and nanobodies offer novel treatment strategies.
• Understanding Trop-2-nanobody interactions is crucial for optimizing therapeutic development.

Purpose of the Study:

• To investigate the molecular interactions between Trop-2 and specific nanobodies (Nb60, Nb65, Nb108).
• To identify key residues and binding poses involved in Trop-2-nanobody interactions.
• To guide the engineering of enhanced Trop-2-targeting nanobodies.

Main Methods:

• Molecular dynamics (MD) simulations combined with MM/GBSA.
• Analysis of binding poses, key residues, and interaction interfaces.
• Experimental validation of engineered nanobody variants.

Main Results:

• MD simulations confirmed binding pose 1 and identified key residues on Trop-2's C-terminal cysteine-poor domain and an α-helix region.
• The length and residues of nanobody CDR3 were critical for Trop-2 binding.
• Engineered nanobody variant Nb-7 showed the highest binding affinity (Kd = 7.931 μM).

Conclusions:

• Molecular dynamics simulations provide valuable insights into Trop-2-nanobody interactions.
• Key interaction sites and CDR3 characteristics were elucidated, guiding nanobody design.
• This research facilitates the development of more effective Trop-2-targeting nanobodies for cancer therapy.