title: Purification of human tissue plasminogen activator with Erythrina trypsin inhibitor.
abstract: Seeds of the legume Erythrina latissima contain a 20,000-dalton, single-chain protein that has been shown to inhibit the amidolytic activity of trypsin and tissue plasminogen activator. It had no comparable effect on urokinase. IC50 values of 1.1 X 10(-7) M for tissue plasminogen activator and 6.9 X 10(-10) M for trypsin were determined by titration. When coupled to agarose, the Erythrina inhibitor provided an effective reagent for affinity purification of tissue plasminogen activator from melanoma cell-conditioned tissue culture medium. Using this as a single-step procedure, 270-fold purified enzyme was reproducibly obtained with yields of 90% or greater. Both one- and two-chain forms of tissue plasminogen activator were purified. The enzyme migrated, in sodium dodecyl sulfate-polyacrylamide gel electrophoresis, as a predominant 72,000-dalton doublet with lesser amounts of immunochemically similar, 115,000- and 68,000-dalton components.
mesh_terms: D002478:Cells, Cultured; D002846:Chromatography, Affinity / Q000379:methods; D004591:Electrophoresis, Polyacrylamide Gel; D007887:Fabaceae; D006801:Humans; D007531:Isoflurophate / Q000494:pharmacology; D008545:Melanoma / Q000201:enzymology; D008970:Molecular Weight; D010946:Plants, Medicinal; D010960:Plasminogen Activators / Q000302:isolation & purification*; D014361:Trypsin Inhibitors / Q000494:pharmacology*; D014568:Urokinase-Type Plasminogen Activator / Q000037:antagonists & inhibitors
chemical_list: D014361:Trypsin Inhibitors; D007531:Isoflurophate; D010960:Plasminogen Activators; D014568:Urokinase-Type Plasminogen Activator