Abstract
Short synthetic oligopeptides based on regions of human proteins that encompass functional motifs are versatile reagents for understanding protein signaling and interactions. They can either mimic or inhibit the parent protein's activity1,2,3,4 and have been used in drug development5. Peptide studies typically either derive peptides from a single identified protein or (at the other extreme) screen random combinatorial peptides4,6, often without knowledge of the signaling pathways targeted. Our objective was to determine whether rational bioinformatic design of oligopeptides specifically targeted to potentially signaling-rich juxtamembrane regions could identify modulators of human platelet function. High-throughput in vitro platelet function assays of palmitylated cell-permeable oligopeptides corresponding to these regions identified many agonists and antagonists of platelet function. Many bioactive peptides were from adhesion molecules, including a specific CD226-derived inhibitor of inside-out platelet signaling. Systematic screens of this nature are highly efficient tools for discovering short signaling motifs in molecular signaling pathways.
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Acknowledgements
The authors thank V. Buckley, Y. Po-Ba, E. Bernard, S. Desgranges and C. Petit for the synthesis of the peptides. This work was funded by Health Research Board, Science Foundation Ireland, and the Programme for Research in Third Level Institutions.
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Contributions
D.C.S., D.K. and N.M. devised the experiment; R.J.E. and D.C.S. formulated the bioinformatics peptide design algorithm and performed the statistical analysis; R.J.E., N.M., D.K. and D.C.S. cowrote the paper; R.J.E. developed the bioinformatics tools and performed the peptide design and the bioinformatics searches for shared or known motifs in bioactive peptides; M.D. designed the selection rules for the synthesis of the peptides to ensure maximum integrity, stability and solubility of the corresponding palmitylated sequences and supervised the synthesis of these peptides; N.M., A.K. and E.D. developed and performed the high-throughput platelet activation assays; G.M. performed the colocalization microscopy; W.S., D.K. and N.M. designed and interpreted the platelet spreading assays; S.D.E.P. performed platelet expression analysis; M.F. provided proteomics data.
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Supplementary information
Supplementary Fig. 1
Schema of method with hypothesized mechanisms of action. (PDF 496 kb)
Supplementary Fig. 2
Example identification of peptide of interest. (PDF 607 kb)
Supplementary Fig. 3
Dose response of CD226_Q palmitylated peptide. (PDF 109 kb)
Supplementary Table 1
Statistical analysis and peptide details for high-throughput assays at two doses. (PDF 48 kb)
Supplementary Table 2
Top 20 motifs shared among peptides identified by SLiMDisc analysis. (PDF 17 kb)
Supplementary Table 3
Potential protein-protein interaction motifs present in active peptides. (PDF 21 kb)
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Edwards, R., Moran, N., Devocelle, M. et al. Bioinformatic discovery of novel bioactive peptides. Nat Chem Biol 3, 108–112 (2007). https://doi.org/10.1038/nchembio854
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DOI: https://doi.org/10.1038/nchembio854
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