Two-step chromatographic purification of recombinant Plasmodium falciparum circumsporozoite protein from Escherichia coli

The Plasmodium falciparum circumsporozoite (PfCS) protein (aa 19–405) has been cloned and expressed in E. coli. The protein was purified in a two-step process that was rapid and reproducible. E. coli cells were grown to a high density before induction for 1 h. Cells were disrupted by high pressure microfluidization and the total bacterial protein solubilized in 6 M Gu-HCl. The protein was refolded while bound to Ni–NTA agarose by exchange of 6 M Gu-HCl for 8 M urea and then slow removal of the urea. The eluted protein was further purified on Q Sepharose Fast Flow using conditions developed to remove E. coli proteins and reduce endotoxin (to 10 EU/50 μg). Yield was 20 mg of PfCS protein from 10 g of wet cell paste. The final protein product bound to HepG2 liver cells in culture and inhibited the invasion of those cells by sporozoites in an ISI assay greater than 80% over control cultures when used at 10 μg/ml.