Self-assembly of heptameric nanoparticles derived from tag-functionalized phi29 connectors.

Abstract

The structure of an induced macromolecular assembly was characterized and found to consist of an ordered heptameric arrangement of recombinant phi29 gp10 connector molecules. Insertion of an N-terminal Strep-II/His(6) tag to the connectors led to the spontaneous formation of large nanoparticles that were distinct from free, wild-type phi29 connectors in both size and symmetry elements. The determination of single-molecule tomograms and image-averaged reconstructions allowed for the stoichiometric and topological characterization of the ordered assemblage, revealing that the nanoparticle is composed of five equatorial connectors arranged with pseudo-5-fold rotational symmetry, capped on its ends by two polar connectors. Additionally, all seven connectors are oriented with their narrower N-terminal necks into the nanoparticle core and wider C-terminal ends out toward the nanoparticle surface, a geometric arrangement accommodated by the shape complementarity of the conical connector profiles. A significant amount of conformational heterogeneity was detected, ranging from changes in overall nanoparticle diameter, to tilting of individual connectors, to variations in connector stoichiometry. Nevertheless, a stable, heptameric nanoparticle was resolved, revealing the significant potential of guided, peptide-mediated supramolecular self-assembly. With this construct, we anticipate the further design of variable N-terminal tags to allow for the generation of nanoparticles with tailored connector stoichiometry and topological arrangements. By modifying the surface-exposed C-terminal ends with application-appropriate moieties, the consistent structure and compact nature of these nanoparticles may prove beneficial in nanotechnological and nanomedical approaches.