Evolution of interleukin-15 for higher E. coli expression and solubility - Inserm - Institut national de la santé et de la recherche médicale Accéder directement au contenu
Article Dans Une Revue Protein Engineering, Design and Selection Année : 2011

Evolution of interleukin-15 for higher E. coli expression and solubility

Résumé

Directed evolution was used to generate IL-15 mutants with increased solubility and cytoplasmic over-expression in Escherichia coli. A protein solubility selection method was used in which the IL-15 gene was expressed as an N-terminal fusion to chloramphenicol acetyltransferase (CAT) as reporter protein. Clones that grew in the presence of high concentrations of chloramphenicol were then screened by ELISA to assay the binding activity of the IL-15-CAT fusion to the IL-15Ra Sushi domain. Two variants of IL-15, M38 and M253, containing five mutations and one mutation respectively, were selected with a dramatic improvement in solubility; the soluble concentration in cell culture was 12-to 18-fold higher, respectively, than for WT IL-15. Characterization of their binding to IL-15Ra and their ability to stimulate the T-cell growth response showed that M38 binds as strongly as native IL-15 to IL-15Ra and acts as an effective agonist of IL-15.
Fichier sous embargo
Fichier sous embargo
Date de visibilité indéterminée
Loading...

Dates et versions

inserm-02442555 , version 1 (16-01-2020)

Identifiants

Citer

Ghislaine Béhar, Véronique Sole, Alain Defontaine, Mike Maillasson, Agnès Quéméner, et al.. Evolution of interleukin-15 for higher E. coli expression and solubility. Protein Engineering, Design and Selection, 2011, 24 (3), pp.283-290. ⟨10.1093/protein/gzq107⟩. ⟨inserm-02442555⟩
43 Consultations
1 Téléchargements

Altmetric

Partager

Gmail Facebook X LinkedIn More