title: Catalytic and fibrinolytic properties of recombinant urokinase plasminogen activator from E. coli, mammalian, and yeast cells.
abstract: The enzymatic and fibrinolytic properties of glycosylated and nonglycosylated recombinant human pro-urokinase (pro-UK) produced in yeast Pichia pastoris were characterized and compared with those of Escherichia coli and mammalian cell-derived pro-UK. Among the five different forms of pro-UK, the yeast glycosylated pro-UK was activated by plasmin with the lowest catalytic efficiency (kcat/Km). The yeast glycosylated urokinase (UK) also had the highest Km in its activation of Glu-plasminogen, and had a substantially lower fibrinolytic activity than the other four forms. These findings suggest that the poly-mannose on Asn-302 of yeast glycosylated pro-UK interfered with its activation by plasmin and its binding interaction with plasminogen. By contrast to plasminogen, the activation of the small synthetic substrate, S2444, was comparable for all five forms of recombinant UK. It is concluded that the glycosyl residue on pro-UK/UK is functionally important and modulates its activatability and its catalytic efficiency against its natural substrate. Therefore, pro-UK from different expression systems cannot be assumed to have comparable fibrinolytic activities.
authors.affiliation: Institute of Molecular Medicine, Nanjing University, 210093, Nanjing, People's Republic of China.
authors.forename: P
authors.initials: P
authors.lastname: Wang
authors.forename: J
authors.initials: J
authors.lastname: Zhang
authors.forename: Z
authors.initials: Z
authors.lastname: Sun
authors.forename: Y
authors.initials: Y
authors.lastname: Chen
authors.forename: V
authors.initials: V
authors.lastname: Gurewich
authors.forename: J N
authors.initials: JN
authors.lastname: Liu
mesh_terms: D000818:Animals; D002384:Catalysis; D005343:Fibrinolytic Agents / Q000494:pharmacology; D006801:Humans; D008322:Mammals; D010960:Plasminogen Activators / Q000235:genetics / Q000494:pharmacology*; D011994:Recombinant Proteins / Q000235:genetics / Q000494:pharmacology*; D012441:Saccharomyces cerevisiae; D013045:Species Specificity; D014568:Urokinase-Type Plasminogen Activator / Q000235:genetics / Q000494:pharmacology*
chemical_list: D005343:Fibrinolytic Agents; D011994:Recombinant Proteins; D010960:Plasminogen Activators; D014568:Urokinase-Type Plasminogen Activator