Self-Assembled Nanobodies as Selectively Targeted, Nanostructured, and Multivalent Materials

Area of Science:

• Biotechnology
• Nanomedicine
• Molecular Engineering

Background:

• Nanobodies are small proteins with therapeutic potential but limited in vivo application due to size and low valency.
• Current limitations restrict nanobodies' capacity for multivalent and cooperative interactions, hindering their effectiveness in complex biological systems.

Purpose of the Study:

• To design modular protein constructs that fuse nanobodies with other protein domains to create self-assembled nanoparticles.
• To enhance nanobody functionality for in vivo applications by increasing valency and promoting stable oligomerization into nanoparticles.
• To maintain nanobody specificity within supramolecular complexes for targeted therapeutic delivery and imaging.

Main Methods:

• Engineered modular protein constructs by fusing nanobodies with additional protein domains.
• Facilitated self-assembly into stable nanoparticles (approx. 70 nm diameter) using C-terminal hexa-histidine tags.
• Utilized nanobodies targeting ricin toxin (A3C8) and Her2 receptor (EM1) as proof-of-concept models.

Main Results:

• Nanobody specificity was preserved in the self-assembled supramolecular complexes.
• A3C8-based nanoparticles efficiently neutralized ricin toxin.
• EM1-based nanoparticles enabled selective imaging of Her2-positive cells, demonstrating dual functionality.

Conclusions:

• Engineered nanobodies into stable, self-assembled nanoparticles (approx. 70 nm) with enhanced multivalent interactivity.
• Demonstrated successful in vivo functionality for both therapeutic neutralization (ricin toxin) and targeted imaging (Her2-positive cells).
• These findings highlight the potential of organized nanobodies in nanoscale assemblies for advanced extracellular and intracellular applications.