Figure 5

Inhibition of thrombin by wtPAI-1, E351A and E351R and Neutralization of PAI-1s by thrombin Thrombin (50 ul, 0.5 U/ml) was incubated with equal amount of wtPAI-1, E351A, or E351R in term of inhibitory activity toward uPA (panel A), or in presence of vitronectin(panel B). After 20, 40, 60 and 120 min, remaining thrombin activities were determined with S-2238. The initial activity of thrombin was taken as 100%. Panel C, Forty nanomolar of thrombin were incubated with 10 nM of active PAI-1 alone, or in presence of 40 nM vitronectin at 37°C. After 30 min, residual PAI-1 activity was determined by titration on uPA. The residual PAI-1 activity without incubation with thrombin was taken as 100%. Empty bars, PAI-1s, filled black bars, PAI-1s + thrombin, and filled light bars, PAI-1s + thrombin + vitronectin.