Crystal structure of Middle East respiratory syndrome coronavirus helicase

Hao, Wei; Wojdyla, Justyna Aleksandra; Zhao, Rong; Han, Ruiyun; Das, Rajat; Zlatev, Ivan; Manoharan, Muthiah; Wang, Meitian; Cui, Sheng

5'-triphosphate DNA (ppp-15T) was synthesized and purified according to previously published procedures [39, 40] . Briefly, ppp-15T oligonucleotide was analyzed by LC/ESI-MS on a XBridge C8 column (2.1 x 50 mm, 2.5μm). Buffer A was 95mM 1,1,1,3,3,3-hexafluoro-2-propanol, 16mM triethylamine in water and buffer B was 100% methanol. A gradient from 2% to 29% B over 26.8 min with flow rate of 0.25 mL/min was employed at a column temperature of 60˚C. A DNAPac-200 column (4 x 250 mm) was used for analytical IE-HPLC. Buffer A was 25mM Tris-HCl, 1mM EDTA in 10% acetronitrile (pH = 8) and buffer B was buffer A plus 1 M sodium bromide. A gradient of 25 to 56% B over 21.5 min at a flow rate of 1.0 mL/min was used at a column temperature of 75˚C. Final purity by IE-HPLC and LC/ESI-MS was above 94% (S3 Fig) .