ENZYMATIC-PROPERTIES OF PHAGE-DISPLAYED FRAGMENTS OF HUMAN PLASMINOGEN

Two low-molecular-mass forms of human plasminogen, plasminogen-(543-79 1)-peptide (micro-plasminogen), comprising the serine protease domain, and plasminogen-(444-791)-peptide (mini-plasminogen), which in additi on contains kringle 5, were displayed on filamentous phage by fusion t o the N-terminus of the minor coat protein pIII, to levels of 0.5 mole cules micro-plasminogen-pIII/phage particle and 0.1 molecules mini-pla sminogen-pIII/phage particle. The proenzymes, quantitatively activated by urokinase, showed catalytic efficiencies that were virtually ident ical to their soluble counterparts, and activity remained associated w ith the phage as demonstrated by phage ELISA and biopanning with human alpha(2)-antiplasmin or the inhibitor Phe-Pro-Arg-CH2Cl. Micro-plasmi nogen-pIII was activated by streptokinase and staphylokinase, two non- enzymatic plasminogen activators, to the same extent as by urokinase. Activated forms of mini-plasminogen-pIII, micro-plasminogen-pIII and m ini-plasminogen dissolved I-125-labelled fibrin films in a dose-depend ent time-dependent manner, with 50% lysis in 20 h requiring 0.52, 3.2 and 0.46 nM active plasmin, respectively. Thus, proenzyme moieties der ived from plasminogen can be successfully displayed on phage with main tenance of their enzymatic properties. The microplasminogen and mini-p lasminogen phage-display systems may be useful to study mechanisms of plasminogen activation.