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

Fui Chin Chong, Wen Siang Tan, Dayang Radiah Awang Biak, Tau Chuan Ling, Beng Ti Tey

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20 Citations (Scopus)

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.

Original languageEnglish
Pages (from-to)1561-1567
Number of pages7
JournalJournal of Chromatography B: Analytical Technologies in the Biomedical and Life Sciences
Volume877
Issue number14-15
DOIs
Publication statusPublished - 15 May 2009
Externally publishedYes

Keywords

  • Escherichia coli
  • Immobilized metal affinity chromatography
  • Nipah virus
  • Nucleocapsid protein

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