Spatial orientation of the alpha and beta sub(c) receptor chain binding sites on monomeric human interleukin-5 constructs

• Published in: The Journal of biological chemistry Vol. 272; no. 33; pp. 20611 - 20618
• Main Authors: Edgerton, MD, Graber, P, Willard, D, Consler, T, McKinnon, M, Uings, I, Arod, CY, Borlat, F, Fish, R, Peitsch, M C, Wells, TNC, Proudfoot, AEI
• Format: Journal Article
• Language: English
• Published: 01.08.1997
• ISSN: 0021-9258

Interleukin-5 (IL-5), a disulfide-linked homodimer, can be induced to fold as a biological active monomer by extending the loop between its third and fourth helices. We have designed eight monomeric IL-5 proteins to optimize biological activity and stability of the monomer. This was achieved by (i) inserting the joining loop at three different positions, (ii) by introducing an additional intramolecular disulfide bridge onto these backbones, and (iii) by creating circular permutations to fix the position of the carboxyl-terminal helix relative to the three other helices. The proteins dimerize with K sub(d) values ranging from 20 to 200 mu M and are therefore monomeric at the picomolar concentrations where they are biologically active. Introduction of a second disulfide confers increased stability, but this increased rigidity results in lower activity of the protein. Contrary to wild type IL-5, mutation of the beta sub(c) contact residue on the first helix, Glu super(12), to Lys, into the circularly permutated constructs, did not abolish TF-1 proliferative and eosinophil activation activities. These results indicate that activation of the IL-5 receptor complex is not mediated solely by Glu super(12) on the first helix, and alternative mechanisms are discussed.