Enhanced periplasmic expression of human activin A in Escherichia coli using a modified signal peptide.
Prep Biochem Biotechnol
; 50(2): 141-147, 2020.
Article
en En
| MEDLINE
| ID: mdl-31647371
Activin A is a member of the transforming growth factor-beta (TGF-ß) protein superfamily, which acts as a hormone in regulating cell proliferation and differentiation. Structurally, activin is a dimer of two subunits linked by a disulfide bond. Since the correct folding of this protein is essential for its function, we aimed to use a modified signal peptide to target the expressed recombinant protein to the periplasm of Escherichia coli as an effective strategy to produce correctly-folded activin A. Therefore, the coding sequence of native Iranian Bacillus licheniformis α-amylase signal peptide was modified and its efficiency was checked by SignalP bioinformatics tool. Then its final sequence was cloned upstream of the activin A mature cDNA. Protein expression was done using 1 mM of isopropyl thio-ß-D-galactoside (IPTG) and a post-induction time of 8 hr. Additionally, following purification of recombinant activin A, circular dichroism spectroscopy was used to determine the accuracy of secondary structure of the protein. Importantly, differentiation of K562 cells to the red blood cell was confirmed by measuring the amount of Fe+2 ions after treatment with recombinant activin A. The results indicated that the produced recombinant activin A had the same secondary structure as the commercial human activin A and was fully functional.
Palabras clave
Texto completo:
1
Colección:
01-internacional
Base de datos:
MEDLINE
Asunto principal:
Señales de Clasificación de Proteína
/
Periplasma
/
Activinas
/
Escherichia coli
Límite:
Humans
Idioma:
En
Revista:
Prep Biochem Biotechnol
Asunto de la revista:
BIOQUIMICA
/
BIOTECNOLOGIA
Año:
2020
Tipo del documento:
Article
Pais de publicación:
Reino Unido