Hr2 406

All crystallization trials were performed at 20 °C using the sitting-drop vapor diffusion method in 96-well sitting-drop vapor diffusion crystallography plates (Art Robbins Instruments, Sunnyvale, CA, USA). Over 600 different conditions from sparse-matrix screening solution kits were tested to identify optimal crystallization conditions. Rod-shaped ScPSAT crystals were obtained over a week from 9.2% (v/v) Tacsimate^TM (pH 5.0) and 16.5% (w/v) PEG 3350 using sparse-matrix screening. To improve the crystals, additional screening was performed using additive (HR2-428, Hampton Research, Viejo, CA, USA) and detergent (HR2-406, Hampton Research, USA) screening. The optimized crystals were obtained from 2.4 M ammonium sulfate and 0.1 M citric acid (pH 3.5), and the average size of the crystals was 200 × 50 × 50 μm.

All crystallization experiments were performed at 20 °C using the sitting-drop vapor diffusion method in 96-well sitting-drop plates (Art Robbins Instruments, Sunnyvale, CA, USA). Approximately 600 different conditions from sparse-matrix screening solution kits were tested to identify the optimal crystallization conditions. The following kits were used: PEG/Ion (HR2-126 and −098), Index (HR2-144), Salt Rx 1/2 (HR2-107 and -109), and Crystal Screen 1/2 (HR2-110 and -112) from Hampton Research (Viejo, CA, USA), Wizard 1/2 (CS-311, Jena Bioscience, Germany), and SG1 Screen (MD1-88, Molecular Dimensions, Rotherham, UK). KlSpdS crystals grew within 24 h in drops containing equal volumes (1 μL) of protein sample (10 mg/mL in 150 mM NaCl, 2 mM DTT, and 20 mM Tris, pH 7.5) and reservoir solution (9.2% v/v TacsimateTM pH 5.0, 16.5% w/v PEG 3350). Additional screening was performed using additive (HR2-428, Hampton Research) and detergent (HR2-406, Hampton Research) screening kits. The optimal crystallization conditions used 9.2% v/v TacsimateTM (pH 5.0), 16.5% (w/v) PEG 3350, and 2.5% (v/v) 1-butanol.