Two distinct forms of plasminogen activator inhibitor in human epidermis and their interaction with urokinase

Abstract

Two species of plasminogen activator (PA) inhibitors (PA-I₁ and PA-I₂) were purified from human cornified cell extract by use of Fast Protein Liquid Chromatographic columns, Mono Q (high performance anion-exchanger) and Mono P (high resolution chromatofocusing). Both PA-I₁ with a molecular weight (MW) of 66, 000 and PA-I₂ with MW 45, 000 showed inhibition against urokinase (UK). However, different inhibition mechanisms were demonstrated for these inhibitors. E-I complex between PA-I₁ and UK was found to be dissociable by SDS-polyacrylamide gel electrophoresis. Whereas, PA-I₂ formed undissociated complex with UK. Using affinity column technique with immobilized active site modified UK, involvement of active site serine for E-I complex formation was investigated. PA-I₁ demonstrated binding affinity with native UK, but it was not adsorbed either to anhydro-UK whose active site serine was converted to dehydroalanine, or DIP-UK whose active site was covered with diisopropylphosphate group. PA-I₂ was found to bind with anhydro-UK in addtion to native UK, but PA-I₂ did not show binding affinity with DIP-UK.