The presence of high and low molecular weight forms of factor VIII in heparinized plasma

Abstract

Low molecular weight Factor VIII is thought to exist either as an integral part of a high molecular weight glycoprotein or as a noncovalently linked component which is dissociated from the large parent molecule in the presence of high calcium concentrations. However, all of this previous work on the molecular structure of Factor VIII has been done using blood which was collected into an anticoagulant that acts by chelating calcium. In order to study the structure of Factor VIII under conditions where the calcium concentrations are maintained at physiological levels during the entire period of collection, blood was taken directly into heparin. This anticoagulant inhibits clot formation by inactivating thrombin but leaves the ionic composition of the plasma unaltered. Under these conditions, more than 50% of the Factor VIII activity is associated with a low molecular weight fraction. This low molecular weight fraction may be effectively separated from the high molecular weight glycoprotein by either gel chromatography or sucrose gradient ultracentrifugation. Further chromatography on a calibrated Sephadex G-100 column shows that the procoagulant molecule has a molecular weight of approximately 20,000. The data support the “two molecule” theory for the structure of Factor VIII and suggest that the many inconsistencies in the literature reflect differences in the ionic environment of the molecule.