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Design of Affinity Chromatography Peptide Ligands Through Combinatorial Peptide Library Screening

Part of the Methods in Molecular Biology book series (MIMB,volume 2178)

Abstract

In this chapter, a protocol to design affinity chromatography matrices with short peptide ligands immobilized for protein purification is described. The first step consists of the synthesis of a combinatorial peptide library on the hydroxymethylbenzoyl (HMBA)-ChemMatrix resin by the divide–couple–recombine (DCR) method using the Fmoc chemistry. Next, the library is screened with the protein of interest labeled with a fluorescent dye or biotin. Subsequently, peptides contained on positive beads are identified by tandem matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS/MS), and those sequences showing greater consensus are synthesized in larger quantities and immobilized on chromatographic supports. Finally, target protein adsorption on peptide affinity matrices is evaluated through equilibrium adsorption isotherms and breakthrough curves.

Key words

  • Peptide ligand
  • Solid-phase peptide synthesis
  • One-bead one-peptide
  • Combinatorial libraries
  • ChemMatrix resin
  • Tandem-mass spectrometry

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  • DOI: 10.1007/978-1-0716-0775-6_16
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Acknowledgments

Work in the author’s laboratories was partially supported by grants from the Universidad de Buenos Aires (UBACyT, 20020170100030BA, PIDAE28), the Agencia Nacional de Promoción Científica y Tecnológica (ANPCyT, PICT-2018-00498), and Consejo Nacional de Investigaciones Científicas y Técnicas de la República Argentina (PIP 11220130100119CO, PU-E 2018 ). M.C. Martínez Ceron, S.L. Giudicessi, M.M. Marani, O. Cascone, and S.A. Camperi are career researchers of the Consejo Nacional de Investigaciones Científicas y Técnicas de la República Argentina (CONICET). 

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Correspondence to S. A. Camperi .

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Barredo, G.R. et al. (2021). Design of Affinity Chromatography Peptide Ligands Through Combinatorial Peptide Library Screening. In: Labrou, N.E. (eds) Protein Downstream Processing. Methods in Molecular Biology, vol 2178. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-0775-6_16

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  • DOI: https://doi.org/10.1007/978-1-0716-0775-6_16

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