Engineering Two-Dimensional Nanobody-Origami Architectures for Enhanced Antiviral Activity

The nanoscale organization of binding ligands offers a powerful strategy to combat viral infections. Herein, we report a programmable 2D DNA origami platform that enables nanoscale control of multivalent nanobody (Nb) spatial patterns for a broad-spectrum antiviral application. By site-selective Nb-DNA conjugation and spatial positioning on 2D origami, we synthesized origami-Nb nanoarchitectures with Nb patterns rationally designed to approximately match the geometric presentation of viral surface proteins. We demonstrate that Nb configurations greatly enhanced viral binding affinity and neutralization potency. For SARS-CoV-2, a triangular Nb pattern matching that of the spike proteins achieved an IC₅₀ of 1.52 nM, representing a 171-fold improvement over monomeric Nbs. Utilizing a gp120-binding Nb, we observed a 233-fold increase in HIV-1 neutralization efficiency using an optimal Nb pattern. The study demonstrates a generalizable strategy for engineering potent viral inhibitors through spatially optimized Nb patterns for high affinity binding of corresponding viral surface antigens.