Production of a biologically active human interleukin 18 requires its prior synthesis as pro-IL-18

Interleukin (IL-)18 is an activator of NK cells and a co-inducer of Th-1 cy tokines, sharing structural features with the IL-1 family of proteins. Unli ke most other cytokines, IL-18 and IL-1 beta lack a signal peptide, have an all beta -pleated sheet structure and are synthesized as biologically inac tive precursors (pro-IL-18 and pro-IL-1 beta), These precursors are cleaved by caspase-1 (IL-1 beta -converting enzyme, ICE) to form the biologically active mature cytokines, Direct expression of mature recombinant human IL-1 8 in E, coli resulted in a partially active cytokine, We tested the possibi lity that correct folding of huIL-18 requires its prior synthesis as pro-IL -18, Because caspase-1 is not readily available, we constructed an expressi on vector encoding human pro-IL-18 in which the caspase-1 cleavage site was mutated into a factor Xa site. To facilitate purification, the mutated pro -IL-18 cDNA was fused in frame to a glutathione-S-transferase (GST) coding sequence. The GST-pro-IL-18 fusion protein was expressed in E, coli, captur ed on glutathione agarose and mature human IL-18, exhibiting high biologica l activity was released upon cleavage with factor Xa, This result indicates that correct folding of huIL-18 occurs at the level of pro-IL-18 and provi des a practical way to produce biologically active huIL-18, (C) 2000 Academ ic Press.