Resistance-based directed evolution of nanobodies for higher affinity in prokaryotes.
Biochim Biophys Acta Gen Subj
; 1868(11): 130710, 2024 Sep 06.
Article
em En
| MEDLINE
| ID: mdl-39245149
ABSTRACT
A prokaryotic resistance-based directed evolution system leveraging protein-fragment complementation assay (PCA) was devised, and its proficiency in detecting protein-protein interactions and discriminating varying degrees of binding affinity was demonstrated by two well-characterized protein pairs. Furthermore, we constructed a random mutant library based on the GBPR36K/E45K mutant, characterized by almost no affinity towards EGFP. This library was subjected to PCA-based prokaryotic directed evolution, resulting in the isolation of back-mutated variants. In summary, we have established an expedited, cost-effective, and structural information-independent PCA-based prokaryotic directed evolution platform for nanobody affinity maturation, featuring tunable screening stringency via modulation of antibiotic concentrations.
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Idioma:
En
Revista:
Biochim Biophys Acta Gen Subj
/
Biochim. biophys. acta, Gen. subj
/
Biochimica et biophysica acta. G, General subjects (Print)
Ano de publicação:
2024
Tipo de documento:
Article
País de publicação:
Holanda