Highly sulfated dermatan sulfate from the skin of the ray Raja montagui: anticoagulant activity and mechanism of action

Abstract

The dermatan sulfate (DS) isolated from the ray skin Raja montagui was identified and characterized. Its average molecular weight (Mw) and sulfate content were 39 kDa and 25% w/w, respectively. This DS prolonged thrombin time and activated partial thromboplastin time and inhibited the thrombin generation in a concentration-dependent manner whereas it had no effect on the anti-Xa assay and on platelet function. Data from the anti-IIa assay allowed the assessment of the specific anticoagulant activity which was 40 units/mg. The kinetics of the thrombin inhibition by heparin cofactor II (HCII) has been studied as a function of DS concentration according to a kinetic model in which the polysaccharide binds quickly to the inhibitor and forms a complex more reactive than the free inhibitor towards thrombin. This DS accelerated thrombin inhibition exclusively by HCII. The dissociation constant of the DS-HCII complex, K(DSHCII), and the rate constant of the thrombin inhibition by this complex, k, were (2.93+/-0.25)x10(-6)M and (2.2+/-0.35)x10(9)M(-1)min(-1), respectively. Our findings indicated that the major polysaccharide in the skin of the ray Raja montagui was a DS endowed with a high anticoagulant effect mediated by HCII and which may constitute an anticoagulant drug of interest in anticoagulant therapy.