Open AccessIdentification of ciliary neurotrophic factor (CNTF) residues essential for leukemia inhibitory factor receptor binding and generation of CNTF receptor antagonists.
Author(s) -
Annalise Di Marco,
Isabelle Gloaguen,
Rita Graziani,
Giacomo Paonessa,
Isabella Saggio,
Keith Hudson,
Ralph Laufer
Publication year - 1996
Publication title -
proceedings of the national academy of sciences of the united states of america
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 5.011
H-Index - 771
eISSN - 1091-6490
pISSN - 0027-8424
DOI - 10.1073/pnas.93.17.9247
Subject(s) - leukemia inhibitory factor receptor , ciliary neurotrophic factor , glycoprotein 130 , alanine scanning , leukemia inhibitory factor , receptor , alanine , biology , biochemistry , microbiology and biotechnology , amino acid , chemistry , signal transduction , mutagenesis , cytokine , neurotrophic factors , interleukin 6 , mutation , genetics , gene , stat3
Ciliary neurotrophic factor (CNTF) drives the sequential assembly of a receptor complex containing the ligand-specific alpha-receptor subunit (CNTFR alpha) and the signal transducers gp130 and leukemia inhibitory factor receptor-beta (LIFR). The D1 structural motif, located at the beginning of the D-helix of human CNTF, contains two amino acid residues, F152 and K155, which are conserved among all cytokines that signal through LIFR. The functional importance of these residues was assessed by alanine mutagenesis. Substitution of either F152 or K155 with alanine was found to specifically inhibit cytokine interaction with LIFR without affecting binding to CNTFR alpha or gp130. The resulting variants behaved as partial agonists with varying degrees of residual bioactivity in different cell-based assays. Simultaneous alanine substitution of both F152 and K155 totally abolished biological activity. Combining these mutations with amino acid substitutions in the D-helix, which enhance binding affinity for the CNTFR alpha, gave rise to a potent competitive CNTF receptor antagonist. This protein constitutes a new tool for studies of CNTF function in normal physiology and disease.