Expression and characterization of soluble human erythropoietin receptor made in Streptomyces lividans 66.
Protein Expr Purif
; 11(3): 271-8, 1997 Dec.
Article
en En
| MEDLINE
| ID: mdl-9425631
A gene encoding the extracellular domain of the human erythropoietin receptor (EPO-R) was constructed using oligonucleotides, with a view to maintaining preferred codon usage for the Streptomycetes. The gene was subcloned into a multicopy Streptomyces-Escherichia coli shuttle vector, pCAN46 (derived from pIJ680), containing a strong constitutive promoter from the S. fradiae aph gene, a signal peptide coding region derived from the protease B gene of S. griseus, and a transcription terminator sequence also derived from the S. fradiae aph gene. Extracellular expression of authentic EPO-R by S. lividans was demonstrated using SDS-PAGE and Western blot analysis, followed by direct amino terminal sequencing of the purified product. Specific binding of S. lividans-expressed EPO-R to recombinant human glycosylated EPO was demonstrated using BIAcore (surface plasmon resonance) analysis and native gel shift assays.
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Colección:
01-internacional
Base de datos:
MEDLINE
Asunto principal:
Receptores de Eritropoyetina
/
Genes Sintéticos
Límite:
Humans
Idioma:
En
Revista:
Protein Expr Purif
Asunto de la revista:
BIOLOGIA MOLECULAR
Año:
1997
Tipo del documento:
Article
País de afiliación:
Canadá
Pais de publicación:
Estados Unidos