Multivalent Ligand–Protein Interactions in DNA‐Confined Compartments: Role of Geometric Match and Electrostatic Screening
TL;DR
It is shown that three‐dimensional nanoscale confinement introduces geometric and electrostatic constraints that critically shape multivalent ligand–protein recognition, providing additional design principles for engineering high‐affinity interactions within synthetic nanocompartments.
Abstract
Multivalent interactions are widely used to enhance ligand binding to selected proteins, yet their occurrence in spatially confined nanoscale environments has been poorly investigated. Here, we use programable DNA origami nanocompartments to study multivalent interactions within nanosized volumes. In our system, encapsulation of oligomeric proteins is mediated by multiple and identical ligands linked to the inner surface of a DNA shell at precisely defined positions. By examining the effect of compartment size and shape as well as ligand spacing and linker length, we identify geometric and electrostatic contributions to the binding affinity and reveal their impact on multivalency. We show that the encapsulation efficiency can be maximized by adjusting linker length, while preserving ligand density and spatial arrangement. Furthermore, electrostatic interactions at the DNA–protein interface can be fine‐tuned to enable sufficient molecular encounters, preventing nonspecific aggregation. Using charged fluorophores to screen the protein surface, binding affinities can be improved by several‐fold as compared to values reported in the literature for similar monovalent ligand–protein interactions. Together, these results show that three‐dimensional nanoscale confinement introduces geometric and electrostatic constraints that critically shape multivalent ligand–protein recognition, providing additional design principles for engineering high‐affinity interactions within synthetic nanocompartments.