Molecular structural studies of human factor VIII.
Neither normal nor hemophilic factor VIII protein enters a 5% sosium dodecyl sulfate gel; on reduction, however, a single 195 000-molecular-weight peptide is observed. Hemophilic and normal factor VIII contain carbohydrate and appear identical in subunit molecular weight, electrical charge, and major antigenic determinants. Thrombin activation and inactivation of factor VIII does not detectably change the subunit molecular weight. Trypsin causes similar activity changes and obviously cleaves the factor VIII subunit. Human plasmin destroys factor VIII procoagulant activity and degrades the factor VIII subunit to 103 000-, 88 000-, and 17 000-molecular-weight peptides. Both normal and hemophilic factor VIII as well as thrombin-inactivated factor VIII support ristocetin-induced platelet aggregation. Purified factor VIII chromatographed on 4% agarose in 1.0 M sodium chloride shows no dissociation of the procoagulant activity from the void volume protein. Gel chromatography on 4% agarose in 0.25 M calcium chloride results in a procoagulant activity peak removed from the void volume protein; both peaks contain protein which does not enter a 5% SDS gel, but on reduction a 195 000-molecular-weight subunit band is observed for each. Both the void volume protein peak and the procoagulant activity peak from the 0.25 M calcium chloride-agarose gel column support ristocetin-induced platelet aggregation. After removal of calcium, a small amount of procoagulant activity is present only in the void volume peak. These data suggest that both the procoagulant and von Willebrand activities are on the same molecule. Thus our previous conclusion remains the same: human factor VIII is a large glycoprotein composed of identical 195 000-molecular-weight subunits jointed by disulfide bonds and is responsible for both antihemophilic and von Willebrand activities in human plasma.
D000327:Adsorption; D000818:Animals; D000906:Antibodies; D001546:Bentonite; D001780:Blood Coagulation Tests; D002848:Chromatography, DEAE-Cellulose; D002850:Chromatography, Gel; D005169:Factor VIII; D005341:Fibrinolysin / Q000494:pharmacology; D006467:Hemophilia A / Q000097:blood*; D006801:Humans; D005779:Immunodiffusion; D008970:Molecular Weight; D010447:Peptide Hydrolases / Q000494:pharmacology; D010973:Platelet Adhesiveness; D010974:Platelet Aggregation; D011487:Protein Conformation; D011817:Rabbits / Q000276:immunology; D013917:Thrombin / Q000494:pharmacology; D014357:Trypsin / Q000494:pharmacology
D000906:Antibodies; D001546:Bentonite; D005169:Factor VIII; D010447:Peptide Hydrolases; D014357:Trypsin; D013917:Thrombin; D005341:Fibrinolysin