Activation of heparin cofactor II by heparin oligosaccharides

Heparin was partially depolymerized with heparinase or nitrous acid. The resulting oligosaccharides were fractionated by gel filtration chromatography and tested for the ability to stimulate inhibition of thrombin by purified heparin cofactor II or antithrombin. Oligosaccharides containing greater than or equal to 18 monosaccharide units were active with antithrombin, while larger oligosaccharides were required for activity with heparin cofactor II. Intact heparin molecules fractionated on a column of immobilized antithrombin were also tested for activity with both inhibitors. The relative specific activities of the unbound heparin molecules were 0.06 with antithrombin and 0.76 with heparin cofactor II in comparison to unfractionated heparin (specific activity = 1.00). We conclude that heparin molecules much greater than 18 monosaccharide units in length are required for activity with heparin cofactor II and that the high-affinity antithrombin-binding structure of heparin is not required.