Improvement of soluble expression of GM-CSF in the cytoplasm of Escherichia coli using chemical and molecular chaperones.

Autor: Malekian R; Department of Pharmaceutical Biotechnology, Isfahan Pharmaceutical Research Center, Faculty of Pharmacy, Isfahan University of Medical Sciences, Isfahan, Iran., Sima S; Department of Pharmaceutical Biotechnology, Isfahan Pharmaceutical Research Center, Faculty of Pharmacy, Isfahan University of Medical Sciences, Isfahan, Iran., Jahanian-Najafabadi A; Department of Pharmaceutical Biotechnology, Isfahan Pharmaceutical Research Center, Faculty of Pharmacy, Isfahan University of Medical Sciences, Isfahan, Iran., Moazen F; Department of Pharmaceutical Biotechnology, Isfahan Pharmaceutical Research Center, Faculty of Pharmacy, Isfahan University of Medical Sciences, Isfahan, Iran., Akbari V; Department of Pharmaceutical Biotechnology, Isfahan Pharmaceutical Research Center, Faculty of Pharmacy, Isfahan University of Medical Sciences, Isfahan, Iran. Electronic address: v_akbari@pharm.mui.ac.ir.
Jazyk: angličtina
Zdroj: Protein expression and purification [Protein Expr Purif] 2019 Aug; Vol. 160, pp. 66-72. Date of Electronic Publication: 2019 Apr 15.
DOI: 10.1016/j.pep.2019.04.002
Abstrakt: The most common approaches to improve soluble expression of heterologous proteins are applications of molecular chaperones such as DnaK, DnaJ, GrpE, GroEL and GroES. The aim of present study was to enhance soluble expression of granulocyte-macrophage colony-stimulating factor (GM-CSF) in Escherichia coli by different approaches including modification of cultivation and induction conditions, and thermally, genetically and chemically enhancement of expression of cellular chaperones. To genetically enhance amount of molecular chaperones, co-expression of pET28-GM-CSF and pKJE7 plasmids was performed. The soluble expressed protein was affinity purified and subjected to endotoxin removal. Co-expression with molecular chaperones significantly increased soluble expression of GM-CSF. Addition of chemical chaperones and osmolytes like NaCl (0.5 M), sucrose (0.5 M), sorbitol (0.5 M) and MgCl 2 (1 mM) to growing media could improve solubility of GM-CSF. Biological activity of purified GM-CSF was confirmed based on its proliferative effect on HL-60 cell lines. The approach developed in the present study can be applied to improve soluble expression of other recombinant protein proteins.
(Copyright © 2019. Published by Elsevier Inc.)
Databáze: MEDLINE
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