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Antitumor activity of interleukin‐21 prepared by novel refolding procedure from inclusion bodies expressed in Escherichia coli
Author(s) -
Asano Ryutaro,
Kudo Toshio,
Makabe Koki,
Tsumoto Kouhei,
Kumagai Izumi
Publication year - 2002
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/s0014-5793(02)03254-4
Subject(s) - escherichia coli , inclusion bodies , inclusion (mineral) , chemistry , microbiology and biotechnology , biochemistry , biology , gene , mineralogy
Interleukin‐21 (IL‐21) has recently been identified as a novel 4‐helix‐bundle type I cytokine possessing a cytokine receptor γ chain essential for the immune response. We report the preparation and functional characterization of Escherichia coli ‐expressed recombinant human IL‐21 (rIL‐21). The rIL‐21, expressed as insoluble inclusion bodies in E. coli , was solubilized and then refolded by using a modified dialysis method. The introduction of redox reagents during refolding led to a dramatic increase in the refolding efficiency. Circular dichroism spectrum analysis showed that the refolded rIL‐21 had an α‐helix as a secondary structure, which is a characteristic of type I cytokines. Flow cytometry confirmed previous reports that rIL‐21 binds to CD3‐activated T cells (T‐LAK) and to cell lines Raji, HL60, and Jurkat. rIL‐21 stimulated the proliferation of T‐LAK but not peripheral blood mononuclear cells, and this effect seems to be identical to that of co‐stimulation with anti‐CD3 antibody. Growth inhibition assay indicated that enhancement of the cytotoxicity of T‐LAK to the human bile duct carcinoma TFK‐1 depended on the concentration of rIL‐21. Thus, refolded rIL‐21 had activity identical to that of authentic IL‐21 and enhanced the anti‐tumor activity of T‐LAK. These conclusions suggest the potential use of the refolded cytokine in adoptive immunotherapy using T‐LAK cells and in the discovery of other functions of the cytokine.