Isolation of tissue-type plasminogen activator-inhibitor complexes from human plasma. Evidence for a rapid plasminogen activator inhibitor.
Plasminogen activator-inhibitor complexes were analyzed by SDS-polyacrylamide gel electrophoresis and enzymography. The complexes appeared as fibrinolytically active bands in the fibrin-indicator gel. A high-molecular-weight t-PA form comigrating with a t-PA-inhibitor complex (Mr 95 000-135 000) from cultured human endothelial cells was purified from plasma by immunoadsorption on anti-t-PA-Sepharose followed by gel filtration on Sephadex G-150. The high-molecular-weight t-PA form was fibrinolytically inactive when assayed by the fibrin-plate method. It was converted to a form with the same electrophoretic mobility as t-PA (Mr 72 000) when treated with 1.5 M NH4OH/39 mM SDS. These observations suggested that the plasma high-molecular-weight t-PA form was an enzyme-inhibitor complex. The complex did not show immunological cross-reactivity with a number of known plasma serine proteinase inhibitors. Both t-PA and u-PA rapidly formed complexes with an inhibitor which was present in plasma in pmolar concentrations. p-Aminobenzamidine blocked the reaction, indicating that the active center of the activator was indeed implicated in complex formation. The complex between the plasma inhibitor and t-PA and the high-molecular-weight t-PA had the same electrophoretic mobilities. The rapid plasminogen activator inhibitor in plasma showed remarkable similarity to a plasminogen activator inhibitor from cultured human endothelial cells. In addition to the high-molecular-weight t-PA form described above, three other t-PA forms were isolated from plasma. Our results indicated that they represented free t-PA and t-PA in complex with respectively C1-esterase inhibitor and alpha 2-antiplasmin.
D002850:Chromatography, Gel; D004591:Electrophoresis, Polyacrylamide Gel; D006023:Glycoproteins / Q000302:isolation & purification*; D006801:Humans; D008970:Molecular Weight; D010960:Plasminogen Activators / Q000037:antagonists & inhibitors*; D015849:Plasminogen Inactivators; D012967:Sodium Dodecyl Sulfate
D006023:Glycoproteins; D015849:Plasminogen Inactivators; D012967:Sodium Dodecyl Sulfate; D010960:Plasminogen Activators