New Insights on Plasmin Long Term Stability and the Mechanism of Its Activity Inhibition Analyzed by Quartz Crystal Microbalance.

We used the research quartz crystal microbalance (RQCM) to monitor regulatory effects of plasmin and trypsin in the presence of their inhibitor α 2 -antiplasmin. The gold surface of quartz crystals was modified with a β-casein layer that served as a substrate for protease digestion. The addition of plasmin or trypsin as well as their mixtures with α 2 -antiplasmin resulted in an increase of resonant frequency, f, and in a decrease of motional resistance, R m , depending on the molar ratio of protease: antiplasmin. At equimolar concentrations of protease and α 2 -antiplasmin (5 nM:5 nM) full inhibition of protease activity took place. Monitoring of plasmin activity on an hourly and daily basis revealed a prominent effect of autolysis and decrease of plasmin activity in freshly activated samples. The degree of inhibition as well as plasmin half-life (t 1/2 = 2.48 ± 0.28 days) connected with its degradation was determined.