Synthesis of conformationally locked L-iduronic acid derivatives: Direct evidence for a critical role of the skew-boat S-2(0) conformer in the activation of antithrombin by heparin

We have used organic synthesis to understand the role of L-iduronic acid co nformational flexibility in the activation of antithrombin by heparin. Amon g known synthetic analogues of the genuine pentasaccharidic sequence repres enting the antithrombin binding site of heparin, we have selected as a refe rence compound the methylated anti-factor Xa pentasaccharide 1. As in the g enuine original fragment, the single L-iduronic acid moiety of this molecul e exists in water solution as an equilibrium between three conformers C-1(4 ), C-4(1) and S-2(0). We have thus synthesized three analogues of 1, in whi ch the L-iduronic acid unit is locked in one of these three fixed conformat ions. A covalent two atom bridge between carbon atoms two and five of L-idu ronic acid was first introduced to lock the pseudorotational itinerary of t he pyranoid ring around the S-2(0) form. A key compound to achieve this con nection was the D-glucose derivative 5 in which the H-5 hydrogen atom has b een replaced by a vinyl group, which is a progenitor of the carboxylic acid . Selective manipulations of this molecule resulted in the S-2(0)-type pent asaccharide 23. Starting from the D-glucose derivative 28, a covalent two a tom bridge was now built up between carbon atoms three and five to lock the L-iduronic acid moiety around the C-1(4) chair form conformation, and the C-1(4)-type pentasaccharide 43 was synthesized. Finally the L-iduronic acid containing disaccharide 58 which, due to the presence of the methoxymethyl substituent at position five adopts a C-4(1) conformation, was directly us ed to synthesize the C-4(1)-type pentasaccharide 61. The locked pentasaccha ride 23 showed about the same activity as the reference compound 1 in an an tithrombin-mediated anti-Xa assay, whereas the two pentasaccharides 43 and 61 displayed very low activity. These results clearly establish the critica l importance of the S-2(0) conformation of L-iduronic acid in the activatio n of antithrombin by heparin.