Structural variation in the antithrombin III binding site region and its occurrence in heparin from different sources.
A tetrasaccharide possessing a biosynthetically permissible structural variability in and adjacent to the antithrombin III (ATIII) binding site has been isolated from heparin lyase depolymerized bovine lung heparin by using strong anion-exchange high-pressure liquid chromatography (SAX-HPLC). On the basis of two-dimensional 500-MHz 1H NMR experiments, including phase-sensitive correlated spectroscopy (COSY) and rotating frame nuclear Overhauser enhancement spectroscopy (ROESY), and fast-atom bombardment mass spectrometry (FAB-MS), the primary structure of this tetrasaccharide was unambiguously established as delta UAp2S (1----4)-alpha-D-GlcNp2S6S(1----4)-beta-D-GlcAp(1----4)-alph a-D-GlcNp2S3S6S (where delta UA represents 4-deoxy-alpha-L-threo-hex-4-enopyranosyluronic acid). The 1H NMR ROESY experiment proved to be particularly valuable in offering sequence information. Heparins from a variety of species and tissue sources were examined by oligosaccharide mapping using SAX-HPLC and gradient polyacrylamide gel electrophoresis. Two of these heparins are used as anticoagulants; they are porcine intestinal mucosal heparin and bovine lung heparin. The predominant ATIII-binding site in porcine heparin contained an N-acetylated glucosamine residue. We now report the structure of the predominant ATIII-binding site in bovine heparin as----4)-alpha-D-GlcNp2S6S(1----4)-beta-D-GlcAp(1----4)-alph a-D- GlcNp2S3S6S(1----4)-alpha-L-IdoAp2S(1----4)-alpha-D-GlcNp 2S6S(1----. This study shows the presence of one or both types of ATIII-binding-site variants in all of the heparins that were examined.
Loganathan
D
D
Division of Medicinal and Natural Products Chemistry, College of Pharmacy, University of Iowa, Iowa City 52242.
Wang
H M
HM
Mallis
L M
LM
Linhardt
R J
RJ
D000818:Animals; D000990:Antithrombin III / Q000378:metabolism*; D001665:Binding Sites; D002236:Carbohydrate Conformation; D002240:Carbohydrate Sequence; D002417:Cattle; D006493:Heparin / Q000378:metabolism*; D019763:Heparin Lyase; D009682:Magnetic Resonance Spectroscopy; D008969:Molecular Sequence Data; D011133:Polysaccharide-Lyases; D012756:Sheep; D013329:Structure-Activity Relationship; D013552:Swine
D000990:Antithrombin III; D006493:Heparin; D011133:Polysaccharide-Lyases; D019763:Heparin Lyase