Contribution of 3-O- and 6-O-sulfated glucosamine residues in the heparin-induced conformational change in antithrombin III |
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Authors: | D H Atha J C Lormeau M Petitou R D Rosenberg J Choay |
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Institution: | Department of Biology, Massachusetts Institute of Technology, Cambridge 02139. |
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Abstract: | The role of 3-O- and 6-O-sulfated glucosamine residues within the heparin octasaccharide critical for biological activity, iduronic acid----N-acetylglucosamine 6-O-sulfate----glucuronic acid----N-sulfated glucosamine 3,6-di-O-sulfate----iduronic acid 2-O-sulfate----N-sulfated glucosamine 6-O-sulfate----iduronic acid 2-O-sulfate----anhydromannitol 6-O-sulfate, was determined by comparing its ability to bind antithrombin, induce a conformational change in this protease inhibitor as monitored by the enhancement of intrinsic fluorescence, and accelerate (at saturation) the interaction of this protein with human factor Xa. The octasaccharide produced a maximum 48% increase in intrinsic fluorescence at 37 degrees C and a rate of factor Xa inhibition of 6 X 10(5) M-1 s-1 as measured by stopped-flow fluorometry at 25 degrees C. The basal rate of the antithrombin-factor Xa interaction observed in the absence of oligosaccharide was 2 X 10(3) M-1 s-1. The synthetic pentasaccharide, consisting of residues 2-6, produced fluorescence enhancement and rate of inhibition equal to those of the octasaccharide. However, a similar pentasaccharide, identical in all respects except that it lacked the 3-O-sulfate on residue 4, produced less than a 5% fluorescence enhancement and a rate of factor Xa inhibition of 8 X 10(3) M-1 s-1. The tetrasaccharide consisting of residues 2-5 produced a 35% fluorescence enhancement and a rate of factor Xa inhibition of 3 X 10(5) M-1 s-1.(ABSTRACT TRUNCATED AT 250 WORDS) |
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