Production and characterization of a fusion form of hepatitis E virus tORF2 capsid protein in Escherichia coli

Abstract

Hepatitis E virus (HEV) is a nonenveloped virus causing an emerging zoonotic disease posing a severe threat to the public health in the world, especially to pregnant women. In this study, a truncated form (aa 368–606) of the open reading frame 2 of the capsid protein (tORF2-HEV), a major structural protein of HEV, was expressed in Escherichia coli. This work characterizes for the first time, the fused Glutathione-S-Transferase-tagged tORF2 (GST-tORF2) and tORF2-HEV forms in E. coli. The fusion protein was purified by affinity chromatography with a purity higher than 90% and to yield about 27% after thrombin digestion. The purified GST-tORF2 protein was then characterized by western blot, using anti-GST antibodies, and CD spectroscopy. The GST-tORF2 and tORF2-HEV proteins were shown to be efficient to develop an ELISA test to detect anti-HEV IgG in mice sera immunized with a recombinant full length ORF2 protein. Sera showed a significant increase of the absorbance signal at 450 nm, in plate wells coated with a quantity of 0.5, 1 and 2 µg of proteins. ELISA plates coated with the purified GST-tORF2 and tORF2-HEV showed similar response when compared to the HEV ELISA where total insect cell lysate, infected with the recombinant baculovirus expressing full ORF2, was used as positive control.

Keywords:

• CD spectroscopy
• ELISA
• GST-tORF2 fusion protein
• Hepatitis E virus
• tORF2-HEV

Acknowledgments

Special thanks to Vidya A. Arankalle, from the Hepatitis Division, National Institute of Virology, Pune, India, for providing us the ELISA plate coated with Sf9 cell lysate infected with ORF2 recombinant baculovirus and the immunized mice sera used in this study.

Ethical approval

This research does not involve the use of Human Participants and/or Animals.

Disclosure statement

No potential conflict of interest was reported by the author(s).