Protein purification was performed via immobilized metal ion affinity chromatography. The pellets were resuspended in the lysis buffer (0.1 M potassium phosphate, 20 mM imidazole, 300 mM NaCl, 10% glycerol, pH 8). Cells were lysed by ultrasonication using a multiprobe sonicator (QSONICA) for a 10 mins process time with 10 s on and 10 s off, 25% amplitude. Cell debris was removed via centrifugation (48 384 g, 45 min, 4 °C). Protein purification was carried out using gravity flow columns and by binding the protein to Ni-agarose resin (Qiagen), in accordance with the manufacturer's instructions. The supernatant was incubated with the resin at 4 °C for 45 min on a rotating mixer, before loading the column. The column was washed with 10 column volumes of lysis buffer, then eluted with elution buffer (0.1 M potassium phosphate buffer, 300 mM imidazole, 300 mM NaCl, 10 % glycerol, pH 8). The eluted protein was concentrated using a 30 000 MW Vivaspin column (GE Healthcare), before being desalted via a PD-10 column (GE Healthcare) and stored in storage buffer (0.1 M potassium phosphate, 300 M NaCl, 30% glycerol, pH 8). SDS-PAGE analysis confirmed the presence of full-length purified protein (Supplementary Figure ). The concentration of purified protein was determined by Fe(II).CO vs. Fe(II)-difference spectroscopy (Supplementary Figure ) as described by Omura and Sato 78 , using the extinction coefficient of 91 mM -1 cm -1 . CO difference spectroscopy was carried out on an Eppendorf BioSpectrometer. The purified protein was concentrated, aliquoted, flash frozen in liquid nitrogen and stored at -80 °C.