Expression and purification of MERS-CoV envelope protein, an essential viroporin, using the baculovirus expression system

Entedar Alsaadi; Dhafer Alghezi; Ian Jones

doi:10.18502/ijm.v15i1.11926

Expression and purification of MERS-CoV envelope protein, an essential viroporin, using the baculovirus expression system

Iran J Microbiol. 2023 Feb;15(1):121-127. doi: 10.18502/ijm.v15i1.11926.

Authors

Entedar Alsaadi¹, Dhafer Alghezi¹, Ian Jones²

Affiliations

¹ Department of Microbiology, College of Medicine, University of Thi-Qar, Thi-Qar, Iraq.
² Department of Biomedical Sciences, School of Biological Sciences, University of Reading, Reading, United Kingdom.

Abstract

Background and objectives: The causative agent of Middle East Respiratory Syndrome (MERS) is a zoonotic Coronavirus (MERS-CoV) identified in Saudi Arabia in 2012. The envelope (E) protein of MERS-CoV is a small viral protein which plays several essential roles during virus replication. To facilitate study of the structure and function of the E protein, recombinant MERS-CoV E protein was expressed using the baculovirus expression system.

Materials and methods: A recombinant E open reading frame including an 8-histidine tag at the amino terminus was designed and cloned into a baculovirus transfer vector. Following construction of a recombinant virus insect cells were infected and the expression of the E protein assessed by SDS-PAGE and Western blotting.

Results: Recombinant E protein, tagged at the N-terminus with a polyhistidine sequence, with a molecular mass of 10.18 kD was identified by Western blotting with an anti-His antibody. Following large scale infection E protein was released by detergent mediated lysis of infected cells and purified by Immobilized Metal Ion Affinity Chromatography (IMAC).

Conclusion: Purified full length recombinant MERS-CoV E protein can be isolated by IMAC and is suitable for further functional, biophysical or immunological studies.

Keywords: Baculovirus; Coronaviruses; Envelope protein; Immobilised metal-affinity chromatography; Insect cells; MERS-CoV; Middle east respiratory syndrome.