Unparalleled purity after one single purification step
After purification with the Strep-tag® system, there is no need for an additional purification step like size exclusion or ion exchange chromatography.
A 36 kDa enzyme and a mutant, both with C-terminal Strep-tag®II, were expressed in the cytoplasm of E. coli. The crude lysate was chromatographically separated on Strep-Tactin® Sepharose® (5 mg/ml) under gravity flow and physiological conditions (100 mM Tris-Cl, pH 8.0). The purification is documented on a Coomassie stained SDS gel where samples from the crude lysate (lane 1), from the flow through (lane 2), and from the elution with 2.5 mM desthiobiotin (lane 3) had been applied. The wild type enzyme is shown to be over 99 % pure (lane 3 contains 50 µg protein vs 0.5 µg protein per band in the molecular size standard). The mutant - although expressed at low level only - could be obtained at high purity under the same conditions.