Construction and characterization of trifunctional single-chain urokinase-type plasminogen activators.
Two chimeric proteins have been constructed. One consists of four parts: a portion of the low molecular mass single-chain urokinase-type plasminogen activator (scu-PA-32K, residues 144-411), a 15-mer linker sequence, the C-terminal amino-acid sequence (residues 53-65) of hirudin (Hir), and an RGD sequence derived from the leech protein decorsin, i.e. scu-PA(32 k)-linker-Hir (residues 53-65)-RGD peptide. The other comprises two main segments: scu-PA(32 k) and hirudin into which RGDSP is inserted between its residues 33 and 34, i.e. hirudin (residues 1-33)-RGDSP-hirudin (residues 34-65)-scu-PA(32 k). These two chimeric genes were expressed in Escherichia coli, and the products were purified by Zn2+-chelating Sepharose 4B chromatography and benzamidine Sepharose 6B chromatography. Our results suggested that these two chimeric proteins not only had plasminogen-dependent fibrinolytic activity, but also possessed platelet aggregation inhibitory activity and antithrombin activity.
Bi
Qun
Q
College of Life Sciences, Peking University, Beijing, China.
Cen
Xiaodong
X
Huang
Yixiu
Y
Zhu
Shenggeng
S
D000925:Anticoagulants / Q000737:chemistry; D001483:Base Sequence; D002850:Chromatography, Gel; D003001:Cloning, Molecular; D017931:DNA Primers; D004926:Escherichia coli / Q000235:genetics; D005337:Fibrin / Q000378:metabolism; D005342:Fibrinolysis; D005822:Genetic Vectors; D006801:Humans; D010975:Platelet Aggregation Inhibitors / Q000737:chemistry; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D014568:Urokinase-Type Plasminogen Activator / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification
D000925:Anticoagulants; D017931:DNA Primers; D010975:Platelet Aggregation Inhibitors; D011993:Recombinant Fusion Proteins; D005337:Fibrin; D014568:Urokinase-Type Plasminogen Activator