20.500.12556/RUL-145024 Focused peptide library screening as a route to a superior affinity ligand for antibody purification 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. biochemistry biophysics biotechnology chemical biology peptidne knjižnice ELISA imunoglobulini kromatografija analizna kemija true false true Angleški jezik Slovenski jezik Članek v reviji 2023-03-30 11:14:52 2023-03-30 11:14:58 2023-03-31 03:36:29 0000-00-00 00:00:00 2021 0 0 12 str. art. 11650 Vol. 11 2021 0000-00-00 Zaloznikova Objavljeno NiDoloceno 0000-00-00 0000-00-00 0000-00-00 543.544+577.27 2045-2322 10.1038/s41598-021-91208-0 65576707 s41598-021-91208-0_(1).pdf s41598-021-91208-0_(1).pdf 1 86A8105F419A49095116DCCB4BBAF5FE c9350e035faaf214c2171390c650f5662b6d489c77ca5af88b718711e4a9ad71 5d02b0de-cedb-11ed-a314-00155dcfd717 https://repozitorij.uni-lj.si/Dokument.php?lang=slv&id=167645 https://www.nature.com/articles/s41598-021-91208-0 1 3d7cd3ba-cedb-11ed-a314-00155dcfd717 https://repozitorij.uni-lj.si/Dokument.php?lang=slv&id=167644 Fakulteta za farmacijo 0 0 0