Production of recombinant rat interleukin-6 in Escherichia coli using a novel highly efficient expression vector pGEX-3T.
Interleukin-6 (IL-6) is one of the most important mediators of the acute phase reaction in liver. For the production of recombinant rat IL-6 in Escherichia coli, a previously isolated cDNA coding for the rat IL-6 was cloned into the modified novel expression vector pGEX-3T. The IL-6 cDNA was highly expressed as a fusion protein with the glutathione S-transferase (GST) at its C-terminus and rat IL-6 at its N-terminus. The GST-IL-6 fusion protein was controlled by a tac-promoter and could be induced very efficiently by isopropyl-beta-D-thiogalactopyranoside. The synthesized GST-IL-6 fusion protein was insoluble and precipitated intracellularly in E. coli. Using an advanced technique, the insoluble protein was solubilized and purified to homogeneity by affinity chromatography using immobilized glutathione in a one-step procedure.
Frorath
B
B
ELIAS Entwicklungslabor, FRG.
Abney
C C
CC
Berthold
H
H
Scanarini
M
M
Northemann
W
W
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015153:Blotting, Western; D004926:Escherichia coli / Q000378:metabolism; D005822:Genetic Vectors; D005982:Glutathione Transferase / Q000096:biosynthesis; D015850:Interleukin-6 / Q000096:biosynthesis / Q000235:genetics* / Q000302:isolation & purification; D008969:Molecular Sequence Data; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification; D014162:Transfection
D015850:Interleukin-6; D011993:Recombinant Fusion Proteins; D005982:Glutathione Transferase