Focused peptide library screening as a route to a superior affinity ligand for antibody purification.
Sci Rep
; 11(1): 11650, 2021 06 02.
Article
em En
| MEDLINE
| ID: mdl-34079028
ABSTRACT
Affinity chromatography is the linchpin of antibody downstream processing and typically relies on bacterial immunoglobulin (Ig)-binding proteins, epitomized by staphylococcal protein A-based ligands. However, such affinity ligands are fairly costly and suffer from chemical instability, leading to ligand denaturation and leaching from chromatographic support. Innovations in this area are aimed at developing robust and highly selective antibody ligands capable of withstanding harsh column sanitization conditions. We report the development and first-stage characterization of a selective short linear peptide ligand of the IgG Fc region capable of capturing all four IgG subclasses. The ligand was discovered through in vitro directed evolution. A focused phage-display library based on a previously identified peptide lead was subjected to a single-round screen against a pool of human IgG. The hits were identified with next-generation sequencing and ranked according to the enrichment ratio relative to their frequency in the pre-screened library. The top enriched peptide GSYWYNVWF displaying highest affinity for IgG was coupled to bromohydrin-activated agarose beads via a branched linker. The resulting affinity matrix was characterized with a dynamic binding capacity of approx. 43 mg/mL, on par with commercially employed protein A-based resin.
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Imunoglobulina G
/
Fragmentos Fc das Imunoglobulinas
/
Cromatografia de Afinidade
/
Evolução Molecular Direcionada
/
Biblioteca de Peptídeos
Tipo de estudo:
Diagnostic_studies
/
Screening_studies
Limite:
Humans
Idioma:
En
Revista:
Sci Rep
Ano de publicação:
2021
Tipo de documento:
Article
País de afiliação:
Eslovênia