Development and Characterization of Peptide Ligands of Immunoglobulin G Fc Region

Affinity chromatography based on bacterial immunoglobulin (Ig)-binding proteins represents the cornerstone of therapeutic antibody downstream processing. However, there is a pressing need for more robust affinity ligands that would withstand the harsh column sanitization conditions, while still disp...

Full description

Saved in:
Bibliographic Details
Published in:Bioconjugate chemistry 2018-08, Vol.29 (8), p.2763-2775
Main Authors: Kruljec, Nika, Molek, Peter, Hodnik, Vesna, Anderluh, Gregor, Bratkovič, Tomaž
Format: Article
Language:English
Subjects:
Affinity
Affinity chromatography
Alanine
Amino Acid Sequence
Antibodies
Antigen presentation
Bacteriophages - genetics
Beads
Binding
Chromatography, Affinity - methods
Combinatorial analysis
Enzyme-Linked Immunosorbent Assay
Glutamine
Humans
Immunoglobulin Fc Fragments - chemistry
Immunoglobulin Fc Fragments - genetics
Immunoglobulin G
Immunoglobulin G - chemistry
Immunoglobulin G - genetics
Ligands
Molecular biology
Molecules
Mutagenesis
Peptides
Peptides - chemistry
Peptides - metabolism
Phages
Protein A
Proteins
Screens
Staphylococcal Protein A - metabolism
Surface Plasmon Resonance
Citations: Items that this one cites
Items that cite this one
Online Access:Get full text
Tags: Add Tag
No Tags, Be the first to tag this record!
Description
Summary:Affinity chromatography based on bacterial immunoglobulin (Ig)-binding proteins represents the cornerstone of therapeutic antibody downstream processing. However, there is a pressing need for more robust affinity ligands that would withstand the harsh column sanitization conditions, while still displaying high selectivity for antibodies. Here, we report the development of linear peptide IgG ligands, identified from combinatorial phage-display library screens. The lead peptide was shown to compete with staphylococcal protein A for the IgG Fc region. Trimming analysis and alanine scanning revealed the minimal structural requirements of the peptide for Fc binding, and the minimized peptide GSYWYQVWF recognized all human IgG subtypes. Mutation of glutamine located at the nonessential position 6 to aspartate led to the optimized peptide GSYWYDVWF with 18-fold higher affinity (K D app. 0.6 μM) compared to the parent peptide. When coupled to paramagnetic beads or a chromatographic matrix, the optimized ligand was shown to selectively enrich antibodies from complex protein mixtures.
ISSN:1043-1802
1520-4812
DOI:10.1021/acs.bioconjchem.8b00395