Crystallization experiments were performed immediately after protein purification. The IPKI variant was dissolved in 50 mM sodium acetate (pH 4.5) and 150 mM NaCl. Protein crystallization was carried out at 20°C using the sitting-drop vapor diffusion method in 288-well crystallization plates (Swissi 3-wells). Wells were filled using a nanodrop-dispensing robot (Mosquito Crystal, TTP Labtech) and various commercial crystallization kits [JCSG (Molecular Dimensions) and PEGs Suite & PEGs II Suite (Qiagen)]. Reservoir solutions were 40 μl in volume, and crystallization drops were composed of 100 nl of reservoir solution and 100, 200, and 300 nl of protein solution at 5 mg/ml. Crystallization plates were sealed with transparent film after setup of the drops and were transferred to a storage cabinet at 20°C. Crystallization was followed using a Rock Imager system (Formulatrix). The IPKI variant was crystallized in 25% polyethylene glycol 3350, 200 mM CaCl2, and 100 mM tris-HCl (pH 8) in 5 days. A single crystal was fished out, washed in the same solution, and flash-cooled in liquid nitrogen. X-ray diffraction was carried out using synchrotron light on the PROXIMA1 beamline at SOLEIL, Gif-sur-Yvette, France. Three thousand six hundred images were collected over 360° and recorded using an Eiger X Dectris detector, with a sample to detector distance yielding a resolution limit of 2.2 Å.

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