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Design of Affinity Chromatography Peptide Ligands Through Combinatorial Peptide Library Screening

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Protein Downstream Processing

Part of the book series: Methods in Molecular Biology ((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.

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References

  1. Cuatrecasas P, Wilchek M, Anfinsen CB (1968) Selective enzyme purification by affinity chromatography. Proc Natl Acad Sci U S A 61:636–643

    Article  CAS  Google Scholar 

  2. Saavedra SL, Barredo GR, Martínez-Ceron MC et al (2018) Design, synthesis and application of short-peptide and peptidomimetic ligands for affinity chromatography. In: Labrou N (ed) Handbook on protein purification: industry challenges and technological developments. Nova Science Publishers, Inc., New York, NY, pp 19–50

    Google Scholar 

  3. Furka A, Sebestyen F, Asgedom M et al (1991) General method for rapid synthesis of multicomponent peptide mixtures. Int J Peptide Protein Res 37:487–493

    Article  CAS  Google Scholar 

  4. Lam KS, Salmon SE, Hersh EM et al (1991) A new type of synthetic peptide library for identifying ligand-binding activity. Nature 354:82–84

    Article  CAS  Google Scholar 

  5. García-Martin F, Quintanar-Audelo M, García-Ramos Y et al (2006) ChemMatrix, a poly(ethylene glycol)-based support for the solid-phase synthesis of complex peptides. J Comb Chem 8:213–220

    Article  CAS  Google Scholar 

  6. Atherton E, Logan CJ, Sheppard RC (1981) Peptide synthesis. Part 2. Procedures for solid-phase synthesis using Nα-fluorenylmethoxycarbonylamino-acids on polyamide supports. Synthesis of substance P and of acyl carrier protein 65-74 decapeptide. J Chem Soc Perkin Trans 1:538–546

    Article  Google Scholar 

  7. Camperi SA, Marani MM, Iannucci NB et al (2005) An efficient strategy for the preparation of one-bead-one-peptide libraries on a new biocompatible solid support. Tetrahedron Lett 46:1561–1564

    Article  CAS  Google Scholar 

  8. Martinez-Ceron MC, Giudicessi SL, Marani MM et al (2010) Sample preparation for sequencing hits from one-bead-one-peptide combinatorial libraries by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. Anal Biochem 400:295–297

    Article  CAS  Google Scholar 

  9. Bray AM, Valerio RM, Maeji NJ (1993) Cleavage of resin-bound peptide esters with ammonia vapour. Simultaneous multiple synthesis of peptide amides. Tetrahedron Lett 34:4411–4414

    Article  CAS  Google Scholar 

  10. Kaiser E, Colescott RL, Bossinger CD et al (1970) Color test for detection of free terminal amino groups in the solid-phase synthesis of peptides. Anal Biochem 34:595–598

    Article  CAS  Google Scholar 

  11. Christensen T (1979) A qualitative test for monitoring coupling completeness in solid phase peptide synthesis using chloranil. Acta Chem Scand 33:763–766

    Article  Google Scholar 

  12. Thermo Scientific. Instructions. Texas red sulfonyl chloride. https://assets.thermofisher.com/TFS-Assets/LSG/manuals/MAN0011223_TexasRed_Sulfonyl_Chloride_UG.pdf. Accessed 23 Feb 2019

  13. Hermanson GT (2013) Fluorescent probes. In: Hermanson GT (ed) Bioconjugate techniques, 3rd edn. Elsevier, New York, NY, pp 395–463

    Chapter  Google Scholar 

  14. Instruction. EZ-Link™ Sulfo-NHS-Biotin. https://assets.thermofisher.com/TFS-Assets/LSG/manuals/MAN0016134_2161850_EZLinkSulfoNHS_Biotin_UG.pdf. Accessed 23 Feb 2019

  15. Hermanson GT (2013) (Strept)avidin–biotin systems. In: Hermanson GT (ed) Bioconjugate techniques, 3rd edn. Elsevier, New York, NY, pp 465–505

    Chapter  Google Scholar 

  16. Hermanson GT (2013) Immobilization of ligands on chromatography supports. In: Hermanson GT (ed) Bioconjugate techniques, 3rd edn. Elsevier, New York, NY, pp 588–740

    Google Scholar 

  17. Miron T, Wilchek MA (1982) Spectrophotometric assay for soluble and immobilized N-hydroxysuccinimide esters. Anal Biochem 126:433–435

    Article  CAS  Google Scholar 

  18. Mellor SL, Welling DA, Fehrentz JA et al (2000) Synthesis of modified peptides. In: Chan WC, White PD (eds) Fmoc solid phase peptide synthesis: a practical approach. Oxford University Press, New York, NY, pp 137–181

    Google Scholar 

  19. Union Biometrica. http://www.unionbio.com Accessed 16 Oct 2012

  20. Christensen C, Groth T, Schiødt CB et al (2003) Automated sorting of beads from a “one-bead-two-compounds” combinatorial library of metalloproteinase inhibitors. QSAR Comb Sci 22:737–744

    Article  CAS  Google Scholar 

  21. Marani MM, Martínez-Ceron MC, Giudicessi SL et al (2009) Screening of one-bead-one-peptide combinatorial library using red fluorescent dyes. Presence of positive and false positive beads. J Comb Chem 11:146–150

    Article  CAS  Google Scholar 

  22. Chan WC, White PD (2000) Basic procedures. In: Chan WC, White PD (eds) Fmoc solid phase peptide synthesis: a practical approach. Oxford University Press, New York, NY, pp 41–76

    Google Scholar 

  23. Chase HA (1984) Prediction of the performance of preparative affinity chromatography. J Chromatogr 297:179–202

    Article  CAS  Google Scholar 

  24. Bradford MM (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72:248–254

    Article  CAS  Google Scholar 

  25. Guy CA, Fields GB (1997) Trifluoroacetic acid cleavage and deprotection of resin-bound peptides following synthesis by Fmoc chemistry. In: Fields GB (ed) Solid phase peptide synthesis. Methods in enzymology, vol 289. Academic, New York, NY, pp 67–83

    Chapter  Google Scholar 

  26. Kodadek T, Bachhawat-Sikder K (2006) Optimized protocols for the isolation of specific protein-binding peptides or peptoids from combinatorial libraries displayed on beads. Mol BioSyst 2:25–35

    Article  CAS  Google Scholar 

  27. Messing A, Stieber A, Gonatas NK (1985) Resolution of diaminobenzidine for the detection of horseradish peroxidase on surfaces of cultured cells. J Histochem Cytochem 33:837–839

    Article  CAS  Google Scholar 

  28. Marani MM, Oliveira E, Côté S et al (2007) Identification of protein-binding peptides by direct matrix-assisted laser desorption ionization time-of-flight mass spectrometry analysis of peptide beads selected from the screening of one bead-one peptide combinatorial libraries. Anal Biochem 370:215–222

    Article  CAS  Google Scholar 

  29. Chan WC, White PD (2000) Fmoc solid phase peptide synthesis: a practical approach. Oxford University Press, New York, NY

    Google Scholar 

  30. Fields GB (1997) Solid-phase peptide synthesis. Methods in enzymology, vol 289. Academic, New York, NY

    Google Scholar 

  31. El-Faham A, Al Marhoon Z, Abdel-Megeed A et al (2013) OxymaPure/DIC: an efficient reagent for the synthesis of a novel series of 4-[2-(2-acetylaminophenyl)-2-oxo-acetylamino] benzoyl amino acid ester derivatives. Molecules 18:4747–4759

    Article  CAS  Google Scholar 

  32. Gairí M, Lloyd-Williams P, Albericio F et al (1990) Use of BOP reagent for the suppression of diketopiperazine formation in boc/bzl solid-phase peptide synthesis. Tetrahedron Lett 31:7363–7366

    Article  Google Scholar 

  33. Lam KS, Lehman AL, Song A et al (2003) Synthesis and screening of “one-bead one-compound” combinatorial peptide libraries. In: Morales GA, Bunin BA (eds) Combinatorial chemistry, Part B. Methods in enzymology, vol 369. Academic, New York, NY, pp 298–322

    Chapter  Google Scholar 

  34. Lam KS, Lebl M (1997) Synthesis of a one-bead one-compound combinatorial peptide library. In: Cabilly S (ed) Combinatorial peptide library protocols. Methods in molecular biology, Human. Totawa, NJ 87, pp 1-6

    Google Scholar 

  35. Camperi SA, Giudicessi SL, Martínez-Ceron MC et al (2016) Combinatorial library screening coupled to mass spectrometry to identify valuable cyclic peptides. Curr Protoc Chem Biol 8:109–130

    Article  Google Scholar 

  36. Al-Warhia TI, Al-Hazimi HMA, El-Faham A (2012) Recent development in peptide coupling reagents. J Saudi Chem Soc 16:97–116

    Article  CAS  Google Scholar 

  37. Boyle R (1966) The reaction of dimethyl sulfoxide and 5-dimethylaminonapthalene-1-sulfonyl chloride. J Org Chem 31:3880–3882

    Article  CAS  Google Scholar 

  38. Martínez-Ceron MC, Giudicessi SL, Kruszyn JN et al (2016) Two-stage screening of combinatorial peptide libraries. Application to bovine serum albumin ligand selection. Rev Cenic Cienc Biol 46:76–86

    Google Scholar 

  39. Lam KS, Lebl M (1992) Streptavidin and avidin recognize peptide ligands with different motifs. ImmunoMethods 1:11–15

    Article  CAS  Google Scholar 

  40. Axelrod D, Hellen EH, Fulbright RM (1991) Total internal reflection fluorescence. In: Lakowicz JR (ed) Topics in fluorescence spectroscopy, biochemical applications, vol 3. Plenum, New York, NY, pp 289–344

    Chapter  Google Scholar 

  41. Adamson AW, Gast AP (1997) Physical chemistry of surfaces. Wiley, New York, NY

    Google Scholar 

  42. Luo Q, Andrade JD (1998) Cooperative adsorption of proteins onto hydroxyapatite. J Colloid Interface Sci 200:104–113

    Article  CAS  Google Scholar 

  43. Bellot JC, Condoret JS (1993) Modelling of liquid chromatography equilibria. Process Biochem 28:365–376

    Article  CAS  Google Scholar 

  44. Carlsson J, Janson JC, Sparman M (1988) Affinity chromatography. In: Janson JC, Rydén L (eds) Protein purification, 2nd edn. Wiley, New York, NY, pp 375–442

    Google Scholar 

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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). 

Author information

Authors and Affiliations

  1. Cátedra de Biotecnología. Facultad de Farmacia y Bioquímica,, Universidad de Buenos Aires, Buenos Aires, Argentina

    G. R. Barredo, S. L. Saavedra, M. C. Martínez-Ceron, S. L. Giudicessi, O. Cascone & S. A. Camperi

  2. Instituto de Nanobiotecnología (NANOBIOTEC). Facultad de Farmacia y Bioquímica., CONICET-Universidad de Buenos Aires, Buenos Aires, Argentina

    G. R. Barredo, S. L. Saavedra, M. C. Martínez-Ceron, S. L. Giudicessi, O. Cascone & S. A. Camperi

  3. Instituto Patagónico para el Estudio de Ecosistemas Continentales (IPEEC), CONICET, Puerto Madryn, Chubut, Argentina

    M. M. Marani

  4. Peptide Science Laboratory, School of Chemistry and Physics, University of KwaZulu-Natal, Durban, South Africa

    F. Albericio

  5. CIBER-BBN, Networking Centre on Bioengineering, Biomaterials & Nanomedicine, Department of Organic Chemistry, University of Barcelona, Barcelona, Spain

    F. Albericio

Authors
  1. G. R. Barredo
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  2. S. L. Saavedra
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  3. M. C. Martínez-Ceron
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  4. S. L. Giudicessi
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  5. M. M. Marani
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  6. F. Albericio
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  7. O. Cascone
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  8. S. A. Camperi
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Corresponding author

Correspondence to S. A. Camperi .

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Editors and Affiliations

  1. Laboratory of Enzyme Technology, Department of Biotechnology, School of Applied Biology and Biotechnology, Agricultural University of Athens, Athens, Greece

    Nikolaos E. Labrou

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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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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-0774-9

  • Online ISBN: 978-1-0716-0775-6

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