Renaturation of recombinant human pro-urokinase expressed in Escherichia coli.
A synthetic gene encoding human pro-urokinase (pro-UK) with E. coli-favored codon usage was cloned into plasmid pET-3d and expressed in E. coli BL21(DE3) LysS strain. The expressed products, which accumulated as inactive inclusion bodies, were denatured and renatured in vitro. A broad range of parameters such as pH, protein concentration, denaturant concentration, the use of cosolvent polyethylene glycol and presence of basic or acidic amino acid was examined. At optimal renaturation condition, pro-UK activity of more than 1000I.U was obtained from 1 milliliter cell culture.
Hua
Z C
ZC
Pharmaceutic Biotechnology Key Laboratory, Department of Biochemistry, Nanjing University, People's Republic of China.
Dong
C
C
Zhu
D X
DX
D003001:Cloning, Molecular; D004792:Enzyme Precursors / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification; D004926:Escherichia coli / Q000235:genetics; D006801:Humans; D006863:Hydrogen-Ion Concentration; D011092:Polyethylene Glycols; D011487:Protein Conformation; D011489:Protein Denaturation; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D014568:Urokinase-Type Plasminogen Activator / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification
D004792:Enzyme Precursors; D011994:Recombinant Proteins; D011092:Polyethylene Glycols; D014568:Urokinase-Type Plasminogen Activator; C067551:saruplase