Mono- and bidimensional 500 MHz 1H-NMR spectra of a synthetic pentasaccharide corresponding to the binding sequence of heparin to antithrombin-III: Evidence for conformational peculiarity of the sulfated iduronate residue

Abstract

1H-NMR spectra of the synthetic pentasaccharide (N-sulfate-6-O-sulfate-α-D-glucosamine) 1→4 (β-D-glucuronic acid) 1→4 (N-sulfate-3,6-di-O-sulfate-α-D-glucosamine) 1→4 (2-O-sulfate-α-L-iduronic acid) 1→4 (N-sulfate-6-O-sulfate-α-D-glucosamine), corresponding to the active site of heparin for antithrombin (AT-III), have been resolved at 500 MHz and assigned by mono- and bidimensional techniques. Vicinal proton coupling constants of the D-glucosamine residues are similar to those in the regular sequences of heparin, indicating that the 4C 1 conformation of the ring, and preference for the g , g conformation of the sulfated hydroxymethyl groups of these residues are neither affected by the unique 3-O-sulfo group nor by sequence effects. By contrast, an unusually large coupling between H-2 and H-3 of the sulfaced L-iduronic acid residue suggests a greater departure from the 1C 4 conformation of this residue when present in the binding sequence to AT-III than in the regular sequences. Such a departure, leading to different orientation and spacing of essential sulfate groups, may have implications for high-affinity binding to AT-III.