Expression and purification of MERS-CoV envelope protein, an essential viroporin, using the baculovirus expression system
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 facilitat...
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| Format: | Article |
| Language: | English |
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Tehran University of Medical Sciences
2023-02-01
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| Series: | Iranian Journal of Microbiology |
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| Online Access: | https://ijm.tums.ac.ir/index.php/ijm/article/view/3881 |
| _version_ | 1797902850521563136 |
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| author | Entedar Alsaadi Dhafer Alghezi Ian Jones |
| author_facet | Entedar Alsaadi Dhafer Alghezi Ian Jones |
| author_sort | Entedar Alsaadi |
| collection | DOAJ |
| description |
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.
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| first_indexed | 2024-04-10T09:23:52Z |
| format | Article |
| id | doaj.art-f5c1d19064d94eb8bb0910c4c4eef31e |
| institution | Directory Open Access Journal |
| issn | 2008-3289 2008-4447 |
| language | English |
| last_indexed | 2024-04-10T09:23:52Z |
| publishDate | 2023-02-01 |
| publisher | Tehran University of Medical Sciences |
| record_format | Article |
| series | Iranian Journal of Microbiology |
| spelling | doaj.art-f5c1d19064d94eb8bb0910c4c4eef31e2023-02-20T08:49:06ZengTehran University of Medical SciencesIranian Journal of Microbiology2008-32892008-44472023-02-0115110.18502/ijm.v15i1.11926Expression and purification of MERS-CoV envelope protein, an essential viroporin, using the baculovirus expression systemEntedar Alsaadi0Dhafer Alghezi1Ian Jones2Department of Microbiology, College of Medicine, University of Thi-Qar, Thi-Qar, IraqDepartment of Microbiology, College of Medicine, University of Thi-Qar, Thi-Qar, IraqDepartment of Biomedical Sciences, School of Biological Sciences, University of Reading, Reading, United Kingdom 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. https://ijm.tums.ac.ir/index.php/ijm/article/view/3881Middle east respiratory syndrome;Coronaviruses;MERS-CoV;Envelope protein;Baculovirus;Insect cells; |
| spellingShingle | Entedar Alsaadi Dhafer Alghezi Ian Jones Expression and purification of MERS-CoV envelope protein, an essential viroporin, using the baculovirus expression system Iranian Journal of Microbiology Middle east respiratory syndrome; Coronaviruses; MERS-CoV; Envelope protein; Baculovirus; Insect cells; |
| title | Expression and purification of MERS-CoV envelope protein, an essential viroporin, using the baculovirus expression system |
| title_full | Expression and purification of MERS-CoV envelope protein, an essential viroporin, using the baculovirus expression system |
| title_fullStr | Expression and purification of MERS-CoV envelope protein, an essential viroporin, using the baculovirus expression system |
| title_full_unstemmed | Expression and purification of MERS-CoV envelope protein, an essential viroporin, using the baculovirus expression system |
| title_short | Expression and purification of MERS-CoV envelope protein, an essential viroporin, using the baculovirus expression system |
| title_sort | expression and purification of mers cov envelope protein an essential viroporin using the baculovirus expression system |
| topic | Middle east respiratory syndrome; Coronaviruses; MERS-CoV; Envelope protein; Baculovirus; Insect cells; |
| url | https://ijm.tums.ac.ir/index.php/ijm/article/view/3881 |
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