Monovalent phage display of human interleukin (hIL)-6: selection of superbinder variants from a complex molecular repertoire in the hIL-6 D-helix

Andrea Cabibbo, Elisabetta Sporeno, Carlo Toniatti, Sergio Altamura, Rocco Savino, Giacomo Paonessa, Gennaro Ciliberto

Research output: Contribution to journalArticle

Abstract

Phage display of proteins can be used to study ligand-receptor interaction and for the affinity-maturation of binding sites in polypeptide hormones and/or cytokines. We have expressed human interleukin-6 (hIL-6) on M13 phage in a monovalent fashion as a fusion protein with the phage coat protein, pIII. Phage-displayed hIL-6 is correctly folded, as judged by its ability to interact with conformation-specific anti-hIL-6 monoclonal antibodies (mAb) and with the hIL-6 receptor complex in vitro. We set up an experimental protocol for the efficient affinity selection of hIL-6 phage using the extracellular portion of the hIL-6 receptor α (hIL-6Rα) fixed on a solid phase. This system was used to affinity-purify from a library of hIL-6 variants, in which four residues in the predicted D-helix of the cytokine were fully randomized, mutants binding hIL-6Rα with higher efficiency than the wild type. When the best-binder variant Q1751/Q183A was combined with a previously identified superbinder S176R [Savino et al., Proc. Natl. Acad. Sci. 90 (1993) 4067-4071], a triple-substitution mutant Q1751/S176R/Q183A (hIL-6IRA) was obtained with a fivefold increased hIL-6Rα binding and a 2.5-fold enhanced biological activity.

Original languageEnglish
Pages (from-to)41-47
Number of pages7
JournalGene
Volume167
Issue number1-2
DOIs
Publication statusPublished - Dec 29 1995

Fingerprint

Bacteriophages
Interleukin-6
Interleukins
Interleukin-6 Receptors
Bacteriophage M13
Cytokines
Peptide Hormones
Capsid Proteins
Libraries
Proteins
Binding Sites
Monoclonal Antibodies
Ligands

Keywords

  • Cytokine
  • fusion protein
  • mutant library
  • phagemid
  • receptor binding
  • screening
  • super-agonism

ASJC Scopus subject areas

  • Genetics

Cite this

Monovalent phage display of human interleukin (hIL)-6 : selection of superbinder variants from a complex molecular repertoire in the hIL-6 D-helix. / Cabibbo, Andrea; Sporeno, Elisabetta; Toniatti, Carlo; Altamura, Sergio; Savino, Rocco; Paonessa, Giacomo; Ciliberto, Gennaro.

In: Gene, Vol. 167, No. 1-2, 29.12.1995, p. 41-47.

Research output: Contribution to journalArticle

Cabibbo, Andrea ; Sporeno, Elisabetta ; Toniatti, Carlo ; Altamura, Sergio ; Savino, Rocco ; Paonessa, Giacomo ; Ciliberto, Gennaro. / Monovalent phage display of human interleukin (hIL)-6 : selection of superbinder variants from a complex molecular repertoire in the hIL-6 D-helix. In: Gene. 1995 ; Vol. 167, No. 1-2. pp. 41-47.
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