The inhibition of protein−carbohydrate interaction provides a powerful therapeutic strategy for the treatment of myriad human diseases. To date, application of such approaches have been frustrated by the inherent low affinity of carbohydrate ligands for their protein receptors. Because lectins typically exist in multimeric assemblies, a variety of polyvalent saccharide ligands have been prepared in the search for high affinity. The cluster glycoside effect, or the observation of high affinity derived from multivalency in oligosaccharide ligands, apparently represents the best strategy for overcoming the “weak binding” problem. Here we report the synthesis of a series of multivalent dendritic saccharides and a biophysical evaluation of their interaction with the plant lectin concanavalin A. Although a 30-fold enhancement in affinity on a valence-corrected basis is observed by agglutination assay, calorimetric titration of soluble protein with a range of multivalent ligands reveals no enhancement in binding free energies. Rather, IC50 values from agglutination measurements correlate well with entropies of binding. This observation suggests that hemagglutination measures a phenomenon distinct from binding that is typified by a large favorable entropy and an unfavorable enthalpy:  this process is almost certainly aggregation. Supporting this assertion, we report crystal structures of multivalent ligands cross-linking concanavalin A dimers. To the best of our knowledge, these structures are the first reported of their kind. Our results indicate that hemagglutination assays evaluate the ability of ligands to inhibit the formation of cross-linked lattices, a process only tangentially related to reversible ligand binding. Cluster glycoside effects observed in agglutination assays must, therefore, be viewed with caution. Such effects may or may not be relevant to the design of therapeutically useful saccharides.

No abstract

The biological importance of S-nitrosothiols (RSNO) and their role in nitric oxide (NO) transport and regulation are now well established. [1][2][3] However, a concrete mechanistic picture for S-nitrosothiol formation, decomposition, and transport, particularly in vivo, is still lacking despite extensive research in this area. [4][5][6] In part, this is due to only a rudimentary knowledge of the structures and conformations of S-nitrosothiols, and how the observed spectroscopic properties are related to the structural features of these substances. Here we present detailed experimental and theoretical results which establish these characteristics.The stabilities of S-nitrosothiols appear to depend on the structure of the organic substituent. 7 Primary species, such as S-nitroso-N-acetylcysteine, are typically unstable and characterized only spectroscopically, 8 whereas tertiary RSNOs, including that derived from N-acetylpenicillamine (SNAP), have been isolated and are indefinitely stable. 9,10 Alkyl substitution also has an effect on both the NMR and UV-vis spectroscopic properties of S-nitrosothiols. Primary species are orange-red, whereas tertiary compounds are green. Both 1 H and 15 N NMR spectra of RSNO compounds show considerable variability in chemical shift, and several demonstrate broad resonances. The relationship between physicochemical properties and bioactivity is not well understood, although primary S-nitrosothiols typically produce shorter-lived biological effects than do tertiary. 1 S-Nitrosothiols exhibit considerable S-N double bond character: as a result, two geometrical isomers are possible. Three crystal structures of S-nitrosothiols have been reported. The tertiary nitrosothiols S-nitroso-N-acetylpenicillamine (SNAP) and S-nitrosotriphenylmethanethiol both exist with the SNO moiety exclusively in the anti orientation. [9][10][11] In contrast, a recent