Cell pellets were resuspended in GST buffer (50 mM Tris/HCl, 300 mM NaCl, 2 mM fresh DTT, pH 7.5) and pressed through a 1000 µL pipette tip. The suspension was mixed with 2.5 µg mL -1 DNase I, 100 µg mL -1 lysozyme and 1x protease inhibitor mix (SERVA, Heidelberg) and incubated for 30 minutes at 4 °C. The cell lysis was carried out using an LM20 Microfluidizer™ at 20,000 psi and centrifugation (17,000 xg, 30 minutes, 4 °C). The supernatant was transferred on a 2 mL glutathione agarose resin (SERVA, Heidelberg) twice, washed three times with 5 mL GST buffer and incubated for 18 hours at 4 °C with 2.87 mg mL -1 PreScission protease (provided by Bianca Berkenfeld) in 2 mL GST buffer. The protein was eluted with 4 mL GST buffer. The eluate was incubated with a spatula tip of FAD solved in 1 mL GST buffer for 1 hour at 4 °C. The protein solution was concentrated (2,981 xg, 10 minutes, 4 °C) and washed with GST buffer using the Amicon ® Pro Purification System (Merck Millipore) with a 10 kDa cut-off. The protein solution was stored at -80 °C until usage.