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Purpose <p>Selective removal of antibodies from complex biological samples (plasma, cell culture medium) is of high importance. There is an increasing demand for efficient biomimetic ligands that mimic Protein A’s affinity towards antibodies and overcome their limitations. </p> Methods <p>In this study, we have designed 25 peptides based on the interaction between human IgG (hIgG) and Protein A using in silico tools. The peptides, FYRGWV and FYGWV, had displayed the best binding energies of -7.6 and − 7.1 kcal/mol and maintained a stable complex with hIgG for 100 ns (MD simulation studies). A porous aldehyde-functionalized poly(hydroxyethyl methacrylate) (PolyHEMA) monolith was used as the chromatography resin for the peptide’s immobilization via Schiff base chemistry. Both FT-IR spectroscopy and XPS have confirmed the successful immobilization of the peptides on the monolith. </p> Results <p>The peptide load on the monolith was found to be 1.3 mg/g monolith (BCA assay data). In the presence of near-neutral pH and physiological salt concentration buffer (pH 7.4), FYRGWV and FYGWV showed static hIgG adsorption of 100.78 mg/g and 50 mg/g and negligible non-specific adsorption for HSA (~ 2.5 mg/g) and transferrin (~ 4.7 mg/g). The primary driving forces of peptide-hIgG interaction are electrostatic, ionic, and hydrophobic. An FYRGWV-PolyHEMA syringe column of 200 µL column volume was successfully fabricated. </p> Conclusion <p>The captured hIgG was successfully eluted from the column with 85% yield using alkaline elution buffers (10 mM phosphate buffer (PB) + 2 mM KCl + 136 mM NaCl (pH 9) and 0.1 M Tris (pH 9)) in sequential mode. Using the FYRGWV-PolyHEMA syringe, hIgG was selectively captured from human serum plasma.</p>

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Rational Design of Protein A-Inspired Peptides and Their Application to Selectively Separate Immunoglobulin G from Human Plasma

  • 期刊论文

  • 发表日期：

    2026年7月19日

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  • Kasturi Goswami,
  • Kishore K. R. Tetala

摘要

Purpose

Selective removal of antibodies from complex biological samples (plasma, cell culture medium) is of high importance. There is an increasing demand for efficient biomimetic ligands that mimic Protein A’s affinity towards antibodies and overcome their limitations.

Methods

In this study, we have designed 25 peptides based on the interaction between human IgG (hIgG) and Protein A using in silico tools. The peptides, FYRGWV and FYGWV, had displayed the best binding energies of -7.6 and − 7.1 kcal/mol and maintained a stable complex with hIgG for 100 ns (MD simulation studies). A porous aldehyde-functionalized poly(hydroxyethyl methacrylate) (PolyHEMA) monolith was used as the chromatography resin for the peptide’s immobilization via Schiff base chemistry. Both FT-IR spectroscopy and XPS have confirmed the successful immobilization of the peptides on the monolith.

Results

The peptide load on the monolith was found to be 1.3 mg/g monolith (BCA assay data). In the presence of near-neutral pH and physiological salt concentration buffer (pH 7.4), FYRGWV and FYGWV showed static hIgG adsorption of 100.78 mg/g and 50 mg/g and negligible non-specific adsorption for HSA (~ 2.5 mg/g) and transferrin (~ 4.7 mg/g). The primary driving forces of peptide-hIgG interaction are electrostatic, ionic, and hydrophobic. An FYRGWV-PolyHEMA syringe column of 200 µL column volume was successfully fabricated.

Conclusion

The captured hIgG was successfully eluted from the column with 85% yield using alkaline elution buffers (10 mM phosphate buffer (PB) + 2 mM KCl + 136 mM NaCl (pH 9) and 0.1 M Tris (pH 9)) in sequential mode. Using the FYRGWV-PolyHEMA syringe, hIgG was selectively captured from human serum plasma.