Cyclic peptide interleukin 5 antagonists mimic CD turn recognition epitope for receptor α

The cyclic peptide AF17121 (Ac‐VDECWRIIASHTWFCAEE) that inhibits interleukin 5 (IL‐5) function and IL‐5 receptor α‐chain (IL‐5Rα) binding has been derived from recombinant random peptide library screening and follow‐up synthetic variation. To better understand the structural basis of its antagonist...

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Published in:Biopolymers 2004-04, Vol.73 (5), p.556-568
Main Authors: Ruchala, Piotr, Varadi, Gyorgyi, Ishino, Tetsuya, Scibek, Jeffery, Bhattacharya, Madhushree, Urbina, Cecilia, Ryk, Donald Van, Uings, Iain, Chaiken, Irwin
Format: Article
Language:English
Subjects:
Amino Acid Sequence
Binding Sites
Binding, Competitive
biosensor interaction analysis
cytokines
Epitopes
Humans
interleukin 5
Interleukin-5 - antagonists & inhibitors
Molecular Mimicry
Peptides, Cyclic - chemistry
protein mimetics
Protein Structure, Secondary
receptor antagonism
Receptors, Interleukin - chemistry
Receptors, Interleukin-5
Structural Homology, Protein
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Summary:The cyclic peptide AF17121 (Ac‐VDECWRIIASHTWFCAEE) that inhibits interleukin 5 (IL‐5) function and IL‐5 receptor α‐chain (IL‐5Rα) binding has been derived from recombinant random peptide library screening and follow‐up synthetic variation. To better understand the structural basis of its antagonist activity, AF17121 and a series of analogs of the parent peptide were prepared by solid phase peptide synthesis. Sequence variation was focused on the charged residues Asp2, Glu3, Arg6, Glu17, and Glu18. Two of those residues, Glu3 and Arg6, form an EXXR motif that was found to be common among library‐derived IL‐5 antagonists. The E and R in the EXXR motif have a proximity similar to charged residues in a previously identified receptor α binding region, the β‐strand between the C‐ and D‐helices of human IL‐5. Optical biosensor interaction kinetics and cell proliferation assays were used to evaluate the antagonist activities of the purified synthetic peptides, by measuring competition with the highly active single chain IL‐5. Analogs in which acidic residues (Asp2, Glu3, Glu17, and Glu18) were replaced individually by Ala retained substantial competition activity, with multiple replacements in these residues leading to fractional loss of potency at most. In contrast, R6A analogs had strongly reduced competition activity. The results reveal that the arginine residue is crucial for the IL‐5Rα binding of AF17121, while the acidic residues are not essential though likely complex‐stabilizing particularly in the Asp2–Glu3 region. By CD, AF17121 exhibited mostly disordered structure with evidence for a small β‐sheet content, and replacement of the arginine had no influence on the observed secondary structure of the peptides. The dominance of Arg6 in AF17121 activity corresponds to previous findings of dominance of the positive charge balance in the antiparallel β‐sheet of IL‐5 composed of 88EERRR92 in one strand of the CD turn region of IL‐5 and with Arg32 in the neighboring β‐strand. These results argue that AF17121 and related library‐derived peptides function by mimicking the CD turn receptor α recognition epitope in IL‐5 and open the way to small molecule antagonist design. © 2004 Wiley Periodicals, Inc. Biopolymers, 2004
ISSN:0006-3525
1097-0282
DOI:10.1002/bip.20001