2Lomonosov Moscow State University, Faculty of Chemistry, 119991 Moscow, Russia; E-mail: email@example.com
3Belozersky Institute of Physico-Chemical Biology, Lomonosov Moscow State University, 11991 Moscow, Russia; E-mail: firstname.lastname@example.org
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Received July 19, 2017; Revision received September 5, 2017
Influenza A virus nuclear export protein NEP (NS2, 14.4 kDa) plays a key role in various steps of the virus life cycle. Highly purified protein preparations are required for structural and functional studies. In this study, we designed a series of Escherichia coli plasmid constructs for highly efficient expression of the NEP gene under control of the constitutive trp promoter. An efficient method for extraction of NEP from inclusion bodies based on dodecyl sulfate treatment was developed. Preparations of purified NEP with either N- or C-terminal (His)6-tag were obtained using Ni-NTA agarose affinity chromatography with yield of more than 20 mg per liter of culture. According to CD data, the secondary structure of the proteins matched that of natural NEP. A high propensity of NEP to aggregate over a wide range of conditions was observed.
KEY WORDS: influenza A virus, nuclear export protein (NEP), affinity chromatography, protein aggregation