Structure and contribution to the heparin cofactor II-mediated inhibition of thrombin of naturally oversulphated sequences of dermatan sulphate.
Dermatan sulphate (DS) obtained from bovine and pig mucosa and pig skin, and charge-enriched fractions of a selected DS preparation, were characterized in terms of charge density, M(r) and disaccharide composition of chondroitin ABC lyase digests, and by 13C-n.m.r. spectroscopy. Besides the major IdoA-GalNAc4SO3 sequences, all DS preparations contain about 10% disulphated disaccharide sequences (mostly IdoA2SO3-GalNAc4SO3, with minor amounts of IdoA-GalNAc4,6SO3). DS fragments (prepared by radical-catalysed depolymerization of DS and retaining the internal structure of the parent polysaccharide) as well as Smith degraded fragments [SD-DS, obtained by controlled degradation of periodate-oxidized and borohydride-reduced DS (RO-DS)] with the general structure GalNAc4SO3(IdoA2SO3-GalNAc4SO3)n-R (where R is the remnant of a glycol-split uronic acid, and n = 2-3 and 3-4) were characterized by one- and two-dimensional 1H-n.m.r., 13C-n.m.r. and disaccharide composition analysis. In accordance with previous findings [Maimone and Tollefsen (1990) J. Biol. Chem. 265, 18263-18271], only fragments with n > or = 3 significantly enhance the heparin cofactor II-mediated inhibition of thrombin. In natural DS preparations and their fractions, this activity (as well as the antithrombotic activity in an animal model) appears to require IdoA2SO3-containing sequences. The heparin cofactor II activity of DS, RO-DS and SD-DS fragments decreases with decreasing M(r). However, RO-DS fragments are more active than DS fragments of similar M(r), probably because of the extra flexibility endowed by glycol-split IdoA residues.
Mascellani
G
G
Opocrin S.p.A. Research and Development Laboratories, Corlo di Formigine, Modena, Italy.
Liverani
L
L
Bianchini
P
P
Parma
B
B
Torri
G
G
Bisio
A
A
Guerrini
M
M
Casu
B
B
D000818:Animals; D002236:Carbohydrate Conformation; D002240:Carbohydrate Sequence; D002417:Cattle; D002850:Chromatography, Gel; D002851:Chromatography, High Pressure Liquid; D002852:Chromatography, Ion Exchange; D003871:Dermatan Sulfate / Q000737:chemistry* / Q000378:metabolism; D015844:Heparin Cofactor II / Q000378:metabolism*; D009682:Magnetic Resonance Spectroscopy; D008969:Molecular Sequence Data; D013464:Sulfuric Acids / Q000737:chemistry; D013552:Swine; D013917:Thrombin / Q000037:antagonists & inhibitors*
D013464:Sulfuric Acids; D003871:Dermatan Sulfate; D015844:Heparin Cofactor II; D013917:Thrombin