Struktur- und Funktionsuntersuchungen am humanen Interleukin-11-Rezeptorkomplex

Abstract

Interleukin-11 (IL-11) belongs to the interleukin-6 (IL-6)-type subfamily of long-chain helical cytokines, which all share the glycoprotein gp130 as a signal transducing receptor component. IL-11 acts on cells expressing gp130 and the IL-11 receptor (IL-11R) alpha-subunit (IL-11R-alpha). The third extracellular, membran proximal domain of the IL-11R-alpha is able to bind with high affinity to IL-11. Using a three-dimensional model of human IL-11, surface exposed amino acid residues of IL-11 were selected for mutagenesis using analogies to the well-characterized receptor recruitment sites of IL-6, CNTF, and LIF. The respective mutants of human IL-11 were expressed as soluble fusion proteins in bacteria and their biological activities were determined. Several mutants with substantially decreased bioactivity were identified and further analyzed in regard to recruitment of IL-11R-alpha and gp130. The structural epitopes of IL-11 required for the recruitment of the individual receptor subunits (sites 1, 2 and 3) have been defined. By plasmon resonance experiments, it was shown that the antagonistic monoclonal antibodies against IL-11 (H2, H56 and E33) bind to amino acids located at site 2. The study an description of the mutant H182L, which engages IL-11R-alpha more efficiently, offer considerable perspectives for the rational design of IL-11 antagonists and hyperagonists. Furthermore, it was shown that the third extracellular domain of the IL-11R-aplha is able to form a ternary complex with IL-11 and gp130 and that it does not require the presence of the cytokine to bind to one molecule gp130. In the third part, no cleavage of the IL-11R-alpha could be detected on macrophages whereas the shedding of the IL-6 receptor alpha-subunit could be induced with LPS and PMA