Purification and properties of a tissue plasminogen activator from hog kidney.

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Abstract
A procedure was developed for the purification of a tissue plasminogen activator from hog kidney. It involves 6 consecutive steps: extraction of the tissue plasminogen activator from acetone-dried hog kidney with 0.3 M potassium acetate buffer, pH 4.2; ammonium sulfate precipitation; hydrophobic chromatography on n-butyl-Sepharose; affinity chromatography on concanavalin A-Sepharose; gel filtration on hydrophilic vinylmonomer (Toyopearl HW-55); affinity chromatography on fibrin-Sepharose. The purified tissue plasminogen activator had an activity of 13,000 IU/mg protein as assessed by plasminogen activation and had no direct fibrinolytic activity. The apparent MW of the purified tissue plasminogen activator was 60,000 as estimated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Carbohydrate analyses of the purified plasminogen activator preparation showed it to contain 54.0 .mu.g of sialic acid and 175.0 g of hexose/mg protein. As assessed by plasmin-catalyzed hydrolysis of D-Val-Leu-Lys-pNA [p-nitroanilide] the functional activity of the purified tissue plasminogen activator was markedly stimulated by addition of fibrin, whereas the interaction with fibrin had a slightly stimulating effect on the activity of human urokinase. Treatment with concanavalin A or glycosidases resulted in 75 or 70% loss of activity of tissue plasminogen activator, respectively, but had no effect on human urokinase.