Purification of Recombinant GST-E6 and Caspase-8 Proteins

Plasmids carrying E6 and caspase 8 (pGEX-E6 and pTriEx-Caspase 8) were previously constructed [22 (link), 24 (link)]. Expression of GST-E6, GST-Caspase 8 and His₆-Caspase 8 in E. coli and subsequent purification were carried out as previously described [22 (link), 24 (link)]. GST-E6, GST-Caspase 8 and His₆-Caspase 8 proteins were diluted into GST protein buffer (PBS pH 8.0, 5% glycerol, 2 mM DTT) and His protein buffer (20 mM HEPES pH 7.4, 150 mM NaCl, 2 mM KCl, 5% glycerol, 2 mM DTT), respectively. The concentration of the proteins was determined using Coomassie Plus – The Better Bradford Assay Reagent (Thermo Scientific, Waltham, MA, USA). Purity of the isolated proteins was assessed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) separation and Coomassie staining.