An efficient system for production of recombinant urokinase-type plasminogen activator.
A system was developed to produce recombinant urokinase-type plasminogen activator in Escherichia coli. The urokinase-type plasminogen activator was produced with a 6 x His-tag at the C-terminus which was shown to have the same activity, after refolding, as the wild-type protein. Purification of the recombinant protein to homogeneity (95%) was possible by one-step affinity chromatography under denaturing conditions. As a result, proteolysis by bacterial proteases during purification was avoided. A higher refolding efficiency (40%) and a higher total recovery yield (25%) of the recombinant protein were obtained by this method.
Tang
W
W
Vascular Research Laboratory, Beth Israel-Deaconess Medical Center, Harvard Medical School, Boston, Massachusetts 02215, USA.
Sun
Z Y
ZY
Pannell
R
R
Gurewich
V
V
Liu
J N
JN
D002846:Chromatography, Affinity; D003001:Cloning, Molecular; D004926:Escherichia coli; D006801:Humans; D007700:Kinetics; D010957:Plasmids; D011489:Protein Denaturation; D017510:Protein Folding; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000302:isolation & purification; D015183:Restriction Mapping; D016324:Sequence Tagged Sites; D014568:Urokinase-Type Plasminogen Activator / Q000096:biosynthesis* / Q000737:chemistry / Q000302:isolation & purification
D011994:Recombinant Proteins; D014568:Urokinase-Type Plasminogen Activator