Recombinant Protein Expression and Purification.

Cultures of Escherichia coli BL21 were grown to an optical density (OD₆₀₀) of 0.6 before inducing the production of recombinant proteins with 0.1 mM isopropyl-β–thiogalactoside. Cultures were incubated at 16 °C with aeration for 16 h and lysed by sonication, and then target proteins were isolated using HisPur Ni-NTA resin (Thermo Fisher Scientific) according to the manufacturer’s instructions. All protein purification steps were performed at 4 °C.