Article ID Journal Published Year Pages File Type
8359320 Protein Expression and Purification 2018 6 Pages PDF
Abstract
Streptococcus pneumoniae is a major pathogen that causes life-threatening diseases, such as pneumonia, otitis media, bacteremia, and meningitis, worldwide and especially in young children and the elderly. Pneumococcal surface protein A (PspA) is a widely studied candidate protein vaccine that represents a promising replacement for current polysaccharide and polysaccharide-conjugate vaccines. In this study, we describe a simple method to produce PspA of clade 4 from an Escherichia coli expression system using hydroxylapatite and ion-exchange chromatography. Using this method, we successfully expressed soluble PspA4 in 10 L of autoinducing culture medium, with a wet-cell yield of 19 g/L and a final PspA4 concentration of 22.8 mg/L. Additionally, we improved PspA4 purity from 17% to 70% in a single step through the use of hydroxylapatite, resulting in acquisition of recombinant PspA4 (>95% purity) at a final yield of 43% from the starting cell-lysis solution. We subsequently verified the secondary structure molecular weight of recombinant PspA4 by circular dichroism and mass spectrometry, respectively. These results demonstrated a highly efficient method for mass producing PspA4 protein and that can also be applied for purification of PspA proteins from other clades.
Related Topics
Life Sciences Biochemistry, Genetics and Molecular Biology Biochemistry
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