Synthesis of three tethered trisaccharides to probe entropy contributions in carbohydrate-protein interactions

Crystal structure data show that the branched trisaccharide 1 constitutes t he complete antigenic determinant of the Salmonella serogroup B antigen tha t is recognized by monoclonal antibody SE155.4. In an effort to characteriz e the entropic costs associated with immobilization of glycosidic torsional angles in the bound state, three distinct intramolecularly tethered analog ues of this trisaccharide 2-4 have been synthesized. Two trisaccharides are tethered by a methylene acetal via the O-2 position of the 3,6-dideoxy-hex ose, abequose to either O-2 of galactose (2) or O-4 of mannose (3). The thi rd tether, alpha,alpha ' -di-thio-p-xylene, spans the C-6 atoms of the mann ose and galactose residues to create trisaccharide 4. The acetal tethers of 2 and 3 span hydroxyl centers that are known to be involved in intramolecu lar sugar-sugar hydrogens bonds, but both trisaccharides are biologically i nactive due to distorted conformations that cannot be accommodated in the a ntibody binding site. Trisaccharide 4 is active since both hydroxymethyl gr oups of galactose and mannose are solvent exposed in the bound state and th e constrained conformation of 4 is virtually superimposable on the bound co nformation of 1. Despite the retained complimentary and significant reducti on of torsional flexibility, trisaccharide 4 exhibits a DeltaG degrees = -7 .6 kcal mol(-1) compared to DeltaG degrees = -7.1 kcal mol(-1) for 1. The m odest free energy gain for the tethered trisaccharide 4 arises from a small entropy gain (TMS = 0.3 kcal mol(-1)) and an even smaller enthalpic change Delta DeltaH = -0.2 kcal mol(-1)).