Chemical, enzymological and pharmacological equivalence of urokinases isolated from genetically transformed bacteria and human urine.

Low molecular weight urokinase (LUK), which was prepared from E. coli containing a plasmid coding for human pro-urokinase, has an amino acid sequence identical to that of LUK isolated from human urine (uLUK) but lacks the carbohydrate side chain at Asn 144 of the B chain. This chemical difference results in an altered mobility in SDS polyacrylamide gel electrophoresis and an apparently increased specific activity of the E. coli-derived product (cLUK) in diffusion-limited test systems (fibrin agar plate tests). Comparative enzymological investigations in homogeneous phases reveal that the active centers and the substrate recognition sites of cLUK and uLUK are congruent. No significant difference between the enzymes was detectable in the following parameters: Michaelis constants and maximum velocities with the synthetic substrate S-2444; activation rates of human and porcine plasminogen; specificity for ten chromogenic substrates; inhibition constants for the competitive inhibitor benzamidine; inhibition by placental urokinase inhibitor and polyclonal antibodies. Further, cLUK and uLUK dissolved fibrin clots prepared from human plasma in vitro with essentially identical velocities. Both, cLUK and uLUK efficiently lysed injected emboli in rabbits and prevented renal fibrin deposition and death due to endotoxin infusion in rats. It is concluded that cLUK, despite the lack of the carbohydrate side chain, is functionally identical and pharmacologically equivalent to uLUK.