A family of leukemia inhibitory factor-binding peptides that can act as antagonists when conjugated to poly(ethylene glycol).
A panel of six naïve 14-residue random peptide libraries displayed polyvalently on M13 phage was pooled and sorted against human leukemia inhibitory factor (LIF). After four rounds of selection, a single large family of peptides with the consensus sequence XCXXXXG(A/S)(D/E)(W/F)WXCF was found to bind specifically to LIF. Peptides within this family did not bind related members of the interleukin-6 family of cytokines, nor to murine LIF that has 80% sequence identity with human LIF. A representative peptide from this family was synthesized and found to bind to LIF with an affinity of approximately 300 nM. The phage-displayed form of this peptide was able to compete with the LIF receptor alpha chain (LIFR) for binding to LIF; however, the free synthetic peptide was unable to inhibit LIF-LIFR binding or inhibit LIF bioactivity in vitro. Using a panel of human/murine chimeric LIF molecules, the peptide-binding site on LIF was mapped to a groove located between the B and the C helices of the LIF structure, which is distinct from the surfaces involved in binding to receptor. To mimic the effect of the phage particle and convert the free peptide into an antagonist of LIFR binding, a 40 kDa poly(ethylene glycol) (PEG) moiety was conjugated to the synthetic LIF-binding peptide. This PEG-peptide conjugate was found to be both an antagonist of LIF-LIFR binding and of LIF signaling in engineered Ba/F3 cells expressing LIFR and the gp130 coreceptor.
Fairlie
W Douglas
WD
The Walter and Eliza Hall Institute of Medical Research, The Cooperative Research Centre for Cellular Growth Factors, Parkville, Victoria 3050, Australia.
Uboldi
Alessandro D
AD
Hemmings
George J
GJ
Smith
Brian J
BJ
Martin
Helene M
HM
Morgan
Phillip O
PO
Baca
Manuel
M
D000595:Amino Acid Sequence; D000818:Animals; D017104:Bacteriophage M13 / Q000235:genetics; D001665:Binding Sites; D002460:Cell Line; D006801:Humans; D015850:Interleukin-6 / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D053605:Leukemia Inhibitory Factor; D053680:Leukemia Inhibitory Factor Receptor alpha Subunit; D051379:Mice; D008969:Molecular Sequence Data; D019151:Peptide Library; D010449:Peptide Mapping; D010455:Peptides / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology; D011092:Polyethylene Glycols / Q000737:chemistry*; D011485:Protein Binding; D015202:Protein Engineering / Q000379:methods; D018121:Receptors, Cytokine / Q000037:antagonists & inhibitors / Q000378:metabolism; D053606:Receptors, OSM-LIF; D011993:Recombinant Fusion Proteins / Q000138:chemical synthesis / Q000235:genetics / Q000378:metabolism; D014162:Transfection; D000093242:Interleukin-6 Inhibitors
D015850:Interleukin-6; D053605:Leukemia Inhibitory Factor; D053680:Leukemia Inhibitory Factor Receptor alpha Subunit; D019151:Peptide Library; D010455:Peptides; D011092:Polyethylene Glycols; D018121:Receptors, Cytokine; D053606:Receptors, OSM-LIF; D011993:Recombinant Fusion Proteins; D000093242:Interleukin-6 Inhibitors; C507837:LIF protein, human; C507840:LIFR protein, human; C507838:Lif protein, mouse; C507841:Lifr protein, mouse