Yeast expression of the cytokine receptor domain of the soluble interleukin-6 receptor.
The complex of the soluble interleukin-6 receptor (sIL-6R) and IL-6 (IL-6) is a potent agonist on cells expressing the signal transducing protein gp 130. In contrast, IL-6 alone only stimulates cells which express a membrane bound form of the IL-6R and gp 130. The natural occurring sIL-6R is generated by shedding of the membrane receptor and to a lesser extend by alternative splicing. We have inserted the coding sequence of the 323 amino acid residues of the human sIL-6R into an expression/secretion vector suitable for the methylotrophic yeast Pichia pastoris. We obtained, however, no detectable expression and secretion of the recombinant protein. When we used only the coding sequence of the cytokine receptor domain of the sIL-6R for the construction of an expression plasmid, this truncated version of the sIL-6R accumulated in the supernatant to 1-5 mg/l. The protein was purified by a single affinity chromatography step using a monoclonal antibody directed against the human IL-6R. Following the same approach, we expressed a truncated splice variant of the sIL-6R. Both, the secreted truncated sIL-6R and the splice variant showed full agonistic biological activity on human hepatoma cells. The described expression strategy will be useful for large scale production of biologically active sIL-6R and might be adapted for the expression of other members of the hematopoietic cytokine receptor family.
Vollmer
P
P
First Department of Medicine, Johannes Gutenberg University of Mainz, Germany.
Peters
M
M
Ehlers
M
M
Yagame
H
H
Matsuba
T
T
Kondo
M
M
Yasukawa
K
K
Büschenfelde
K H
KH
Rose-John
S
S
D015703:Antigens, CD / Q000096:biosynthesis* / Q000737:chemistry; D005822:Genetic Vectors / Q000378:metabolism*; D010843:Pichia / Q000378:metabolism*; D011487:Protein Conformation; D018121:Receptors, Cytokine / Q000096:biosynthesis*; D018123:Receptors, Interleukin / Q000096:biosynthesis* / Q000737:chemistry; D019947:Receptors, Interleukin-6; D012995:Solubility
D015703:Antigens, CD; D018121:Receptors, Cytokine; D018123:Receptors, Interleukin; D019947:Receptors, Interleukin-6