TXA2 is a potent vasoconstrictor and a powerful inducer of platelet aggregation. Despite an intense interest in TXA2 and its involvement in numerous physiological functions, relatively little is known about the enzyme which controls its production (thromboxane synthase). Thromboxane synthase is a microsomal enzyme that is found in a number of tissues including platelets, lung and brain. The platelet enzyme is thought to play an important role in thrombosis and hemostasis; however, the function of this enzyme in lung and brain and the molecular similarities of the enzyme from different tissues is still unknown. Substantial purification of the enzyme from human platelets and bovine lung has been achieved in our laboratory using affinity chromatography. Affinity columns are made by coupling avidin to CNBr activated Sepharose 4B. The avidin is then used to remove thromboxane synthase that has been reacted with biotin-U63557A (A specific thromboxane synthase inhibitor). The further development of this method is being pursued. In addition we have produced and characterized a monoclonal antibody to thromboxane synthase. The antibody reacts to a single protein band when it is used in a Western blot of an SDS-PAGE electrogram of partially purified human platelet enzyme or bovine brain enzyme. The antibody reacts to two proteins in the partially purified bovine lung enzyme. Since our antibody appears to react with the enzyme from the platelet, lung and brain of several species, we hope to use it in the purification and immunological characterization of the enzyme from various tissues. The proposed research will provide basic biochemical information on the chemical, physical, immunological and kinetic characteristics of thromboxane synthase, thus aiding in the design of specific thromboxane synthase inhibitors.
Showing the most recent 10 out of 63 publications
