title: Purification of histidine-tagged nucleocapsid protein of Nipah virus using immobilized metal affinity chromatography.
abstract: 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 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 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 an inlet N protein concentration of 0.4 mg/ml in a scale-up immobilized metal affinity chromatography (IMAC) packed bed column of Nickel Sepharose 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.
authors.affiliation: Department of Chemical and Natural Resources Engineering, Faculty of Engineering, Universiti Malaysia Pahang, Kuantan, Pahang, Malaysia.
authors.forename: Fui Chin
authors.initials: FC
authors.lastname: Chong
authors.forename: Wen Siang
authors.initials: WS
authors.lastname: Tan
authors.forename: Dayang Radiah Awang
authors.initials: DR
authors.lastname: Biak
authors.forename: Tau Chuan
authors.initials: TC
authors.lastname: Ling
authors.forename: Beng Ti
authors.initials: BT
authors.lastname: Tey
mesh_terms: D002846:Chromatography, Affinity / Q000379:methods*; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D006639:Histidine / Q000235:genetics / Q000378:metabolism; D008670:Metals / Q000737:chemistry*; D045405:Nipah Virus / Q000235:genetics*; D019590:Nucleocapsid Proteins / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism
chemical_list: D008670:Metals; D019590:Nucleocapsid Proteins; D011993:Recombinant Fusion Proteins; D006639:Histidine