3.3. SEN Crystallization

Screening of crystallization condition was performed at 4 °C by using an automated liquid-handling robotic system (Formulatrix, Nano Transfer 8) and crystallization screening kits from Hampton Research (Aliso Viejo, CA, USA) (Crystal Screen Suite, Crystal Screen 2 Suite, PEG/Ion Screen Suite, PEG/Ion 2 Screen Suite, Index Suite, PEGRx 1 Suite, and PEGRx 2 Suite) and Molecular Dimensions (MIDAS Suite, SG1 Screen Suite, PACT premier Suite, Structure Screen 1 Suite, Structure Screen 2 Suite, Morpheus Suite, Stura Footprint Screen Suite, 3D Structure Screen Suite, MultiXtal Suite, MacroSol Suite, JCSG-plus Suite, and ProPlex Suite). Sitting drops (0.5 μL) consisting of 0.25 μL of the protein solution and 0.25 μL of the reservoir solution were equilibrated against 30 μL of the reservoir solution in 96-well MRC plates (Molecular Dimensions, Newmarket, Suffolk, UK). After three days, tiny polycrystals were grown under the condition containing 2% (v/v) Tacsimate™ pH 4.0 and 16% (w/v) polyethylene glycol (PEG) 3350. Diffraction quality crystals were obtained under the optimized conditions containing 2% (v/v) Tacsimate™ pH 4.0 and 12% (w/v) PEG 3350.