Characterization and Carbohydrate Specificity of Pradimicin S

The pradimicin family of antibiotics is attracting attention due to its anti-infective properties and as a model for understanding the requirements for carbohydrate recognition by small molecules. Members of the pradimicin family are unique among natural products in their ability to bind sugars in a Ca2+-dependent manner, but the oligomerization to insoluble aggregates that occurs upon Ca2+ binding has prevented detailed characterization of their carbohydrate specificity and biologically relevant form. Here we take advantage of the water solubility of pradimicin S (PRM-S), a sulfated glucose-containing analogue of pradimicin A (PRM-A), to show by NMR spectroscopy and analytical ultracentrifugation that at biologically relevant concentrations, PRM-S binds Ca2+ to form a tetrameric species that selectively binds and engulfs the trisaccharide Manα1–3(Manα1–6)Man over mannose or mannobiose. In functional HIV-1 entry assays, IC50 values of 2–4 μM for PRM-S corrrelate with the concentrations at which oligomerization occurs as well as the affinities with which PRM-S binds the HIV surface envelope glycoprotein gp120. Together these data reveal the biologically active form of PRM-S, provide an explanation for previous speculations that PRM-A may contain a second mannose binding site, and expand our understanding of the characteristics that can engender a small molecule with the ability to function as a carbohydrate receptor.