Purification and some properties of the protein component of tissue thromboplastin from human brain

Abstract

The protein component of tissue thromboplastib (Factor III) from human brain was purified by extraction of a microsomal fraction with sodium deoxycholate, gel filtration of the extract on Sephadex G-100 and preparative polyacrylamide-gel electrophoresis in the presence of sodium dodecyl sulphate. The product, apoprotein III, was homogeneous by anayltical polyacrylamide-gel electrophoresis, and it induced monospecific antibodies in rabbits and goat as shown by immunodiffusion and immunoelectrophoresis. Amino acid- and carbohydrate-analysis data for apoprotein III are presented. The carbohydrate moiety of the protein consists of fucose, mannose, galactose, N-acetylglucosamine and N-acetylneuraminate, amounting to a total content of 6.3g/100g. The apoprotein alone had no procoagulant activity. When Factor III was reconstituted by combining the pure apoprotein with a purified lipid fraction from the deoxycholate extract of crude Factor III, a high and optimal procoagulant activity was obtained at a phospholipid/protein ratio of 1.1g/g. Phosphatidylethanolamine alone had a weak but significant ability to restore activity, whereas phosphatidylcholine and phosphatidylserine separately had almost none. Two-component mixtures were on average more effective, and three-component mixtures far more effective, than the single phospholipids. The inclusion of a small amount of phosphatidylserine was very important for high activity.

Affinity purification of human tissue factor: interaction of factor VII and tissue factor in detergent micelles.

“…Based on these results, we conclude that we have purified human tissue factor to an essentially homogeneous form, which has an apparent Mr of -46,000 as judged by NaDodSO4/PAGE with and without mercaptoethanol (not shown). This material is significantly different from previously described preparations of human brain tissue factor (19,20) in two important respects. First, it has a lower apparent Mr (46,000 vs. 53,000).…”

Thrombosis-Inducing Activity in Plasma of Patients with Acute Respiratory Tract Infection Disappears after Treatment

“…Tissue factor (coagulation factor III, tissue throm boplastin) is a membrane glycoprotein that serves as the nonenzymatic cofactor for factor VII and repre sents a potent procoagulant activity [17,18]. Signif icantly higher levels of tissue factor activity are ex pressed on the surface of peripheral monocytes isolat ed from patients with infectious diseases when compared with the cells from normal control subjects [19,20].…”

Synthesis of Thromboplastin (Tissue Factor) by Endothelial Cells

“…[1] and shown to be a glycoprotein with an apparent relative molecular weight around 50 kD [1,2]. The yield of our method was too low to permit much protein chemis try with the then available methods.…”

“…A light-chain component must, therefore, be coded for by a separate gene. Evidence from studies of purified apo protein III seems to exclude the necessity of a light chain for TPL acitivty [2].…”