Journal ArticleDOI
Preparation and properties of bovine factor VIII (antihemophilic factor)
Gordon A. Vehar,Earl W. Davie +1 more
TLDR
These experiments involving three highly specific serine proteases support the conclusion that the triplet observed on polyacrylamide gels is factor VIII.Abstract:
Factor VIII has been purified approximately 300000-fold from bovine plasma by ammonium sulfate fractionation, glycine precipitation, DEAE-Sephadex column chromatography, sulfate--Sepharose column chromatography, Sephadex G-200 gel filtration, and factor X--Sepharose column chromatography. The highly purified preparation migrated as a triplet on sodium dodecyl sulfate/urea--polyacrylamide gel electrophoresis with apparent molecular weights of 93000, 88000, and 85000. The coagulant activity of the purified preparations was inhibited by antibodies raised in rabbits against either the purified factor VIII protein or a preparation of factor VIII/von Willebrand factor. Antibodies to the purified protein also inhibited the coagulant activity of factor VIII/von Willebrand factor preparations. The purified factor VIII contained no platelet-aggregating activity, as measured in human platelet-rich plasma. The purified preparation of factor VIII was required for the activation of factor X in the presence of factor IXa, calcium, and phospholipid. It was activated about 30-fold by thrombin or factor Xa plus calcium and phospholipid, and each of these reactions was accompanied by a change in the sodium dodecyl sulfate/urea--polyacrylamide gel electrophoresis pattern of the protein. Factor VIII was rapidly inactivated by bovine-activated protein C in a reaction requiring calcium and phospholipid. This reaction was also associated with a change in the sodium dodecyl sulfate/urea--polyacrylamide gel electrophoresis pattern of the highly purified protein. These experiments involving three highly specific serine proteases support the conclusion that the triplet observed on polyacrylamide gels is factor VIII.read more
Citations
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Journal ArticleDOI
The coagulation cascade: initiation, maintenance, and regulation
Journal ArticleDOI
Deficiency of protein C in congenital thrombotic disease.
TL;DR: It is suggested that the recurrent thrombotic disease in this family is due to an inherited deficiency in protein C, a potent in vitro anticoagulant enzyme and an in vivo profibrinolytic agent.
Journal ArticleDOI
The roles of protein C and thrombomodulin in the regulation of blood coagulation.
Journal ArticleDOI
Molecular cloning of a cDNA encoding human antihaemophilic factor.
John J. Toole,John L. Knopf,John M. Wozney,L A Sultzman,J L Buecker,Debra D. Pittman,Randal J. Kaufman,Eugene L. Brown,Charles B. Shoemaker,Elizabeth C. Orr +9 more
TL;DR: A complete copy of the mRNA sequences encoding human coagulation factor VIII:C has been cloned and expressed and has an obvious domain structure, contains sequence repeats and is structurally related to factor V and ceruloplasmin.
Journal ArticleDOI
Structure of human factor VIII.
Gordon A. Vehar,Bruce Keyt,Dan L. Eaton,Henry Rodriguez,Donogh P. O'Brien,F. Rotblat,Herman Oppermann,Rodney G. Keck,William I. Wood,Richard N. Harkins,Edward G. D. Tuddenham,Richard M. Lawn,Daniel J. Capon +12 more
TL;DR: Determination of the thrombin cleavage sites in plasma-derived factor VIII polypeptides allows prediction of the domains involved in the associated activation and inactivation of the protein.
References
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Journal ArticleDOI
Molecular weight analysis of oligopeptides by electrophoresis in polyacrylamide gel with sodium dodecyl sulfate.
R.T. Swank,K.D. Munkres +1 more
TL;DR: Because of the importance of intrinsic charge and conformation, the system, although allowing a first approximation in molecular weight determination, may also be applicable to peptide “mapping,” particularly for “insoluble” peptide mixtures with prominent hydrophobic association, such as encountered in cellular membranes, viruses, and proteolytic digests.
Journal ArticleDOI
Immunization, isolation of immunoglobulins, estimation of antibody titre.
N. M. G. Harboe,Agnete Ingild +1 more
TL;DR: High titred monospecific antibodies are obtained in rabbits injecting as little as 25 μg pure antigen per kg body weight, and antibody titres as measured by 4 different titration methods are compared using anti‐human IgG from rabbits as a model.
Journal ArticleDOI
Anticoagulant properties of bovine plasma protein C following activation by thrombin.
Journal ArticleDOI
A Brain Extract as a Substitute for Platelet Suspensions in the Thromboplastin Generation Test
W. N. Bell,H. G. Alton +1 more
TL;DR: This work has devised a modification of the Biggs and Douglas test, which it is felt allows a higher degree of accuracy, as well as solving the problem and eliminating the need for siliconized glassware.
Journal ArticleDOI
Quantitative Assay of a Plasma Factor Deficient in von Willebrand's Disease that is Necessary for Platelet Aggregation. RELATIONSHIP TO FACTOR VIII PROCOAGULANT ACTIVITY AND ANTIGEN CONTENT
TL;DR: An assay for the von Willebrand factor (VIII(VWF), based on the observation that a log-log relationship exists between the amount of ristocetin-induced aggregation of washed, normal platelets and the concentration of normal plasma present in the test system, suggests that VIII( VWF) assayed in this study may be the "anti-bleeding factor" that is deficient in vonWillebrand's disease.
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