Preparation of functionally active recombinant human interleukin-6.
The gene of human interleukin-6 (hIL-6) with an additional 20 amino acids on the N-end, including six histidine residues, was cloned into the expression plasmid pET28b(+). The conditions were elaborated for preparing highly active protein both using denaturing agents and without them. Application of a dialysis cascade allowed us to prepare a functionally active hIL-6 of 90-95% purity with the yield of 3 mg from liter of the cell culture. The highest activity was detected by ELISA in the preparation obtained without denaturing agents. The functional activity of hIL-6 was studied by flow cytofluorimetry. Addition of hIL-6 to the cells of immortal lines of human multiple myeloma resulted in dimerization of the gp130 receptor molecule.
Spiridonova
V A
VA
Belozersky Institute of Physico-Chemical Biology, Lomonosov Moscow State University, 119992 Moscow, Russia. spiridon@belozersky.msu.ru
Lygina
A S
AS
Anohina
M M
MM
Tupitsyn
N N
NN
D045744:Cell Line, Tumor; D004797:Enzyme-Linked Immunosorbent Assay; D004926:Escherichia coli / Q000378:metabolism; D006801:Humans; D015850:Interleukin-6 / Q000096:biosynthesis* / Q000302:isolation & purification / Q000494:pharmacology; D011994:Recombinant Proteins / Q000096:biosynthesis
D015850:Interleukin-6; D011994:Recombinant Proteins