We present the bioconjugation of avidin as a central and/or bridging building block with mono-, bi-and tetravalent biotinylated glycodendrimers to fabricate defined supramolecular nanostructures for future (bio)medical applications. For this purpose mono-, bi-and tetravalent biotinylated glycodendrimers, decorated with short alkyl-linked or long PEG-linked biotin ligands, were synthesized and characterized by NMR, IR and mass spectrometry and HABA displacement assay. Various techniques (UV/Vis, DLS, TEM, LILBID-MS and AF4) were used in order to obtain information about the structural properties of different conjugates of avidin and mono-, bi-and tetravalent biotinylated glycodendrimers. The biotin ligand's spacer length, its chemical structure and the degree of biotin functionalization are essential parameters in the formation of nanostructures with avidin having a controlled composition and size dimension up to 100 nm. Biohybrid structures with avidin as a central unit require monovalent glycodendrimers with PEG-linked biotin, while bi-and tetravalent glycodendrimers with short alkyl-linked biotin ligands are more efficient than their counterparts with longer PEG-biotin ligands in the fabrication of defined biohybrid structures (B up to 100 nm) with avidin as a bridging unit. The most dominating key issue, combined with other conjugation issues, is the optimal ligand-receptor stoichiometry to fabricate biohybrid structures with diameter of <20, <30 or up to 100 nm.

Biohybrid structures formed by noncovalent interaction between avidin as a bridging unit and biotinylated glycodendrimers based on poly(propyleneimine) (GD-B) have potential for biomedical application. Therefore, an exact knowledge about molar mass, dispersity, size, shape, and molecular structure is required. Asymmetrical flow field-flow fractionation (AF4) was applied to separate pure and assembled macromolecules according to their diffusion coefficients. The complex biohybrid structures consist of single components (avidin, differently valent GD-B) and nanostructures. These nanostructures were systematically studied depending on the degree of biotinylation and ligand−receptor stoichiometry by AF4 in combination with dynamic and static light scattering detection. This enables the quantification of composition and calculation of molar masses and radii, which were used to analyze scaling properties and apparent density of the formed structures. These data are compared to hydrodynamic radii obtained by applying the retention theory to the AF4 data. It is shown that depending on their architecture the molecular shape of biohybrid structures is changed from rod-like to spherical toward network-like behavior.

A series of adamantyl-modified glycodendrimers (mPPI-Gx-AdaA-C) was prepared in a two-step synthesis using two efficient reactions: (1) urea bond formation from amine and isocyanate and (2) reductive amination. 1H NMR spectroscopy (host guest titration and ROESY experiments) was used to evaluate the graded effect of steric hindrance as a function of the number and type of oligosaccharide molecules and of the number of adamantyl (Ada) units on the complexation with monomeric β-cyclodextrin (β-CD). Glycosylated fourth generation PPIs showing an average substitution in adamantyl groups of 13% were found to interact with β-CD effectively, and were considered as candidates for further complexation studies with a polymeric cyclodextrin derivative (poly-β-CD). The host–guest interaction features of the maltosylated dense shell glycodendrimer along with the low cytotoxicity provided the rational basis for the use of these adamantyl-functionalized glycodendrimers in the design of supramolecular systems potentially useful as healthcare materials.