Separation and purification of periplasmic cytochrome c553 using reversed micelles

Surfactant concentration, ionic strength, and pH were optimised for the sel ective separation and purification of periplasmic cytochrome c553 from reco mbinant E. coil TG2 cells using response surface methodology. Back-extracti on was accomplished using counter-ionic surfactant addition. Optimum forwar d extraction conditions were: 65 mM bis(2-ethylhexyl)sulfosuccinate sodium salt (AOT), 0.07 M NaCl, and pH 8.4, while the optimum back-extraction cond itions were 80 mM trioctylmethylammonium chloride, 0.85 M KCl, and pH 9.62. In comparison to a conventionally purified sample using column chromatogra phy (10 mg cytochrome c553 l(-1) with a purity of 0.66), reverse micelles a chieved the same concentration and similar purity (0.50) in only two simple steps.