title: High-level expression and purification of human pro-UK cDNA in Escherichia coli.
abstract: A chemically synthesized human pro-urokinase (pro-UK) CDNA was cloned into the expression vector PET-11d and expressed in E. coli BL21(DE3) pLysS under the control of the T7 promoter. Using 0.1 mmol/L IPTG induction, the expression level of the recombinant pro-UK attained up to 15% of the total bacterial proteins and existed as inclusion bodies. After denaturation and renaturation in vitro, the expressed pro-UK was purified to be identified by Zn2+ selective precipitation, immuno-affinity chromatography, and benzamidine affinity adsorption. The specific activity of the purified human pro-UK was about 110,000 IU/mg.
authors.affiliation: Department of Biochemistry, State Key Laboratory of Pharmaceutical Biotechnology, Nanjing University, China.
authors.forename: Y
authors.initials: Y
authors.lastname: Xue
authors.forename: G
authors.initials: G
authors.lastname: Peng
authors.forename: Z
authors.initials: Z
authors.lastname: Ma
authors.forename: Y
authors.initials: Y
authors.lastname: Chen
authors.forename: D
authors.initials: D
authors.lastname: Zhu
mesh_terms: D003001:Cloning, Molecular; D018076:DNA, Complementary; D004926:Escherichia coli; D015870:Gene Expression; D005822:Genetic Vectors; D006801:Humans; D002479:Inclusion Bodies; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification; D012967:Sodium Dodecyl Sulfate; D014568:Urokinase-Type Plasminogen Activator / Q000096:biosynthesis / Q000235:genetics* / Q000302:isolation & purification
chemical_list: D018076:DNA, Complementary; D011994:Recombinant Proteins; D012967:Sodium Dodecyl Sulfate; D014568:Urokinase-Type Plasminogen Activator; C067551:saruplase