Plasma PCIs have similar MW (∼ 57K), amino acid composition, and heparin dependence (Suzuki et al 1983, JBC 258:163) as urinary UKI (Stump et al 1986, JBC 261:12759). Urinary PCI of ∼ 50K MW has a similar heparin dependence and urokinase (UK) competes with activated protein C (APC) for this PCI (Geiger et al 1986, Circ. 74:11-234). For comparison, three forms of PCI, one from urine and two from plasma, were purified, and each exhibited heparin-dependent UK and APC inhibitory activity and formed heparin-dependent complexes with APC. The APC-PCI complexes were visible on immunoblots (nondenaturing gels) developed using: A) monoclonal anti-UKI + 125I-antimouse IgG; B) polyclonal anti-plasma PCI + 125I-plasma PCI; and C) monoclonal anti-protein C + 125I-protein C. The three forms of purified PCI were detected by methods A and B. Two new bands of APC-inhibitor complexes were seen upon incubation of plasma with APC in the presence of heparin, and the same pattern was visualized by methods A, B, and C. In the absence of heparin, only one APC-inhibitor band was visualized by methods A and B, but two bands were visualized by method C. Plasma immunodepleted of UKI by monoclonal anti-UKI-Sepharose showed no detectable antigen or complexes with APC as visualized by methods A and B. However, the UKI-depleted plasma contained components which formed a reduced amount of complexes with APC as visualized with protein C antibodies, i.e. method C. Heparin stimulates tenfold the PCI activity of normal plasma. Based on amidolytic assays of APC using S-2366, the UKI-depleted plasma was very deficient (< 15%) in heparin-dependent PCI activity, whereas the weak heparin-independent PCI activity was slightly reduced. This indicates that the majority of heparin-dependent PCI activity of plasma is immunologically.related to UKI. These studies suggest that the two slightly different forms of plasma PCI, the urinary UKI, and the urinary PCI are very similar if not identical proteins and that plasma may contain a minor heparin-independent PCI which is not immunologically related to these proteins.
