Purification of histidine-tagged nucleocapsid protein of Nipah virus using immobilized metal affinity chromatography

Nucleocapsid (N) protein of Nipah Virus (NiV) is a potential serological marker used in the diagnosis of NiV infections. In this Study, a rapid and efficient purification system, HisTrap (TM) 6 Fast Flow packed bed Column was applied to purify recombinant histidine-tagged N protein of NiV from clarified feedstock. The optimizations of binding and elution conditions of N protein of NiV onto and from Nickel Sepharose (TM) 6 Fast Flow were investigated. The optimal binding was achieved at pH 7.5, Superficial velocity of 1.25 cm/min. The bound N protein was successfully recovered by a stepwise elution with different concentration of imidazole (50, 150, 300 and 500 mM). The N protein of NiV was captured and eluted from at inlet N protein concentration of 0.4 mg/ml in a scale-up immobilized metal affinity chromatography (IMAC) packed bed column of Nickel Sepharose (TM) 6 Fast Flow with the optimized condition obtained from the method Scouting. The Purification of histidine-tagged N protein using IMAC packed bed column has resulted a 68.3% yield and a purification factor of 7.94. (C) 2009 Elsevier B.V. All rights reserved.