Saccharide receptor achieves concentration dependent mannoside selectivity through two distinct cooperative binding pathways

Tetrapodal receptor 1 relies upon structural flexibility to reveal new binding modes for saccharide recognition and to achieve unique pyranoside binding affinity and concentration dependent selectivity. The association constants, K_as, between 1 and eight pyranosides commonly found in cell surface glycans were measured in CDCl₃ by ¹H NMR titrations, revealing a preference for α- and β-octyl mannopyranosides (α-Man and β-Man). Whereas most of the pyranosides studied-α/β-octyl glucopyranoside (α/β-Glc), α/β-octyl galactopyranoside (α/β-Gal), and α/β-octyl N-acetylglucosaminopyranoside (α/β-GlcNAc)-bind 1 in a 1 : 1 stoichiometry at 25 °C, β-Man exclusively forms a 2 : 1 receptor-pyranoside complex. Alternatively, in an excess of pyranoside, 1 binds α- and β-Man in a 1 : 2 receptor : pyranoside stoichiometry with a high degree of positive cooperativity (K₂/K₁ ∼ 13.7 and 7.6 for α- and β-Man respectively) and selectivities as high as 16.8 : 1 α-Man : α-Gal. Moreover, this preference changes as a function of pyranoside concentration, favoring β-Glc at low concentration (<0.1 mM) and favoring mannosides at higher concentrations. The thermodynamic binding parameters (ΔH⁰ and ΔS⁰) reveal that the cooperativity in the second binding events drive the formation of 1 ₂:β-Man or 1:β-Man₂ because of a decrease in unfavorable entropy upon each second binding event compared to the first. The structures of the complexes were determined by 1D and 2D ¹H NMR spectroscopy in combination with molecular modeling. The 1:β-Man ₂ complex exhibits C₂ symmetry, where both β-Man equivalents bind identical sites within 1, such that the pyranosides within the complex are symmetrically equivalent. Alternatively, 1₂:β-Man is a cage-like structure where only three of the aminopyrrolitic arms of the receptor are involved in binding, leaving a fourth available for further functionalization in later generation receptors. Multivalency and cooperativity are ubiquitous in Nature, and 1 utilizes these modes of recognition to achieve selectivity for monosaccharide residues.