Purification of Recombinant Proteins

Elute from HisTrap consisting of the respective protein fractions were centrifuged at 39,000 g for 15 min, and the supernatant was loaded on a 5 ml StrepTrap HP column (GE HealthCare). The binding buffer was 150 mM NaCl, 50 mM Tris pH 8.0, and the elution buffer was 150 mM NaCl, 50 mM Tris pH 8.0, 2.5 mM desthiobiotin. Elution buffer was injected into the column in two rounds of 5 ml each (Fraction I and II, respectively). The fractions with the protein were pooled, concentrated, and injected into the Superdex-75 column (GE HealthCare) to elute with a final buffer containing 50 mM NaCl, 50 mM Tris pH 8.0 (A50).
In the case of {AB^ΔCt}, only HisTrap and StrepTrap were performed, followed by washing off desthiobiotin using 50 mM NaCl, 50 mM Tris pH 8.0 (A50) buffer during protein concentration and consequently stored.