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Focused peptide library screening as a route to a superior affinity ligand for antibody purification
ID Bozovičar, Krištof (Avtor), ID Jenko Bizjan, Barbara (Avtor), ID Meden, Anže (Avtor), ID Kovač, Jernej (Avtor), ID Bratkovič, Tomaž (Avtor)

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Izvleček
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.

Jezik:Angleški jezik
Ključne besede:biochemistry, biophysics, biotechnology, chemical biology
Vrsta gradiva:Članek v reviji
Tipologija:1.01 - Izvirni znanstveni članek
Organizacija:FFA - Fakulteta za farmacijo
Status publikacije:Objavljeno
Različica publikacije:Objavljena publikacija
Leto izida:2021
Št. strani:12 str.
Številčenje:Vol. 11, art. 11650
PID:20.500.12556/RUL-145024 Povezava se odpre v novem oknu
UDK:543.544+577.27
ISSN pri članku:2045-2322
DOI:10.1038/s41598-021-91208-0 Povezava se odpre v novem oknu
COBISS.SI-ID:65576707 Povezava se odpre v novem oknu
Datum objave v RUL:30.03.2023
Število ogledov:381
Število prenosov:50
Metapodatki:XML RDF-CHPDL DC-XML DC-RDF
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Gradivo je del revije

Naslov:Scientific reports
Skrajšan naslov:Sci. rep.
Založnik:Nature Publishing Group
ISSN:2045-2322
COBISS.SI-ID:18727432 Povezava se odpre v novem oknu

Licence

Licenca:CC BY 4.0, Creative Commons Priznanje avtorstva 4.0 Mednarodna
Povezava:http://creativecommons.org/licenses/by/4.0/deed.sl
Opis:To je standardna licenca Creative Commons, ki daje uporabnikom največ možnosti za nadaljnjo uporabo dela, pri čemer morajo navesti avtorja.

Sekundarni jezik

Jezik:Slovenski jezik
Ključne besede:peptidne knjižnice, ELISA, imunoglobulini, kromatografija, analizna kemija

Projekti

Financer:ARRS - Agencija za raziskovalno dejavnost Republike Slovenije
Številka projekta:P4-0127
Naslov:Farmacevtska biotehnologija: znanost za zdravje

Financer:Drugi - Drug financer ali več financerjev
Program financ.:University of Ljubljana Innovation Fund
Številka projekta:820-1/2020-33

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