title: Optimization of extracellular truncated staphylococcal protein A expression in Escherichia coli BL21 (DE3).
abstract: Staphylococcal protein A (SpA) plays an important role in Staphylococcus aureus pathogenesis. The recombinant SpA is also widely used in biotechnology to purify polyclonal and monoclonal immunoglobulin G antibodies. In this study, expression and secretion of a truncated form of SpA containing five immunoglobulin-binding domains using its own native signal sequence were optimized in Escherichia coli. Optimization was carried out using response surface method (RSM), making use of the interaction between five variables. The initial results revealed that the signal peptide from S. aureus was recognized in E. coli and the resulting SpA was expressed and secreted into the medium. Compounds, such as glycine, affected the secretion of SpA into the culture medium. The central composite design experiment showed that the optimum conditions for the maximum expression of recombinant truncated SpA in E. coli included 10% (w/v) lactose, 1.77% (w/v) glycine, induction time of 11 H, an optical density (600) of 1.1, and a temperature of 33 °C. Optimization using RSM resulted in a fivefold increase in the secretion of SpA. To date, this is the first study of its kind regarding the definite influence of glycine concentration and duration of the cultivation period on the secretion of SpA.
authors.affiliation: Department of Molecular Genetics, National Institute of Genetic Engineering and Biotechnology (NIGEB), Tehran, Iran.
authors.forename: Garshasb
authors.initials: G
authors.lastname: Rigi
authors.forename: Samira Ghaed
authors.initials: SG
authors.lastname: Mohammadi
authors.forename: Maryam Rezaei
authors.initials: MR
authors.lastname: Arjomand
authors.forename: Gholamreza
authors.initials: G
authors.lastname: Ahmadian
authors.forename: Kambiz Akbari
authors.initials: KA
authors.lastname: Noghabi
mesh_terms: D001709:Biotechnology; D003470:Culture Media; D004926:Escherichia coli / Q000235:genetics*; D015964:Gene Expression Regulation, Bacterial; D011994:Recombinant Proteins / Q000096:biosynthesis; D013205:Staphylococcal Protein A / Q000096:biosynthesis* / Q000302:isolation & purification; D013211:Staphylococcus aureus / Q000235:genetics*
chemical_list: D003470:Culture Media; D011994:Recombinant Proteins; D013205:Staphylococcal Protein A
keywords: CCD; E. coli; RSM; SpA; optimization; secretion