Affinity Chromatography of Insulin with a Heptapeptide Ligand Selected from Phage Display Library

Abstract

A heptapeptide phage display library was screened with insulin to find its ligands for affinity chromatography. The peptide was synthesized and coupled to EAH Sepharose 4B (5.4 μmol mL−1 bed). Then, insulin chromatography was carried out with mobile phases of different pH values and by the addition of urea and ethylene glycol. It was found that electrostatic interactions were predominant for the affinity binding, and hydrogen bonding might also contribute somewhat to the affinity. Finally, frontal analysis was performed and the dynamic binding capacity of the affinity column for insulin at 50% breakthrough was estimated at 60.6 mg mL−1 bed, which was about two times higher than the theoretical binding capacity of the monomeric insulin. The result suggests that insulin was bound in dimer state in a stoichiometric relationship with the coupled peptide, indicating the high binding efficiency of the peptide ligand for insulin.

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Acknowledgments.

This work was financially supported by the Natural Science Foundation of China (grant 20025617).

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Correspondence to Yan Sun.

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Yu, HQ., Dong, XY. & Sun, Y. Affinity Chromatography of Insulin with a Heptapeptide Ligand Selected from Phage Display Library. Chromatographia 60, 379–383 (2004). https://doi.org/10.1365/s10337-004-0407-5

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Keywords

  • Column liquid chromatography
  • Affinity chromatography
  • Peptide ligand
  • Phage display library
  • Insulin