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Purification and Functional Characterization of a Biologically Active Full-Length Feline Immunodeficiency Virus (FIV) Pr50Gag

1
Department of Microbiology & Immunology, College of Medicine and Health Sciences (CMHS), United Arab Emirates University (UAEU), Al Ain 2000, UAE
2
Department of Anatomy, College of Medicine and Health Sciences (CMHS), United Arab Emirates University (UAEU), Al Ain 20000, UAE
3
Department of Biochemistry, College of Medicine and Health Sciences (CMHS), United Arab Emirates University (UAEU), Al Ain 20000, UAE
4
Université de Strasbourg, CNRS, Architecture et Réactivité de l’ARN, UPR 9002 Strasbourg, France
*
Author to whom correspondence should be addressed.
These authors contributed equally to this work.
Viruses 2019, 11(8), 689; https://doi.org/10.3390/v11080689
Received: 4 July 2019 / Revised: 23 July 2019 / Accepted: 25 July 2019 / Published: 27 July 2019
(This article belongs to the Section Animal Viruses)
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Abstract

The feline immunodeficiency virus (FIV) full-length Pr50Gag precursor is a key player in the assembly of new viral particles. It is also a critical component of the efficient selection and packaging of two copies of genomic RNA (gRNA) into the newly formed virus particles from a wide pool of cellular and spliced viral RNA. To understand the molecular mechanisms involved during FIV gRNA packaging, we expressed the His6-tagged and untagged recombinant FIV Pr50Gag protein both in eukaryotic and prokaryotic cells. The recombinant Pr50Gag-His6-tag fusion protein was purified from soluble fractions of prokaryotic cultures using immobilized metal affinity chromatography (IMAC). This purified protein was able to assemble in vitro into virus-like particles (VLPs), indicating that it preserved its ability to oligomerize/multimerize. Furthermore, VLPs formed in eukaryotic cells by the FIV full-length Pr50Gag both in the presence and absence of His6-tag could package FIV sub-genomic RNA to similar levels, suggesting that the biological activity of the recombinant full-length Pr50Gag fusion protein was retained in the presence of His6-tag at the carboxy terminus. Successful expression and purification of a biologically active, recombinant full-length Pr50Gag-His6-tag fusion protein will allow study of the intricate RNA-protein interactions involved during FIV gRNA encapsidation. View Full-Text
Keywords: feline immunodeficiency virus (FIV); retroviral RNA packaging; viral assembly; Pr50Gag protein expression; Gag protein purification; His-tag fusion protein feline immunodeficiency virus (FIV); retroviral RNA packaging; viral assembly; Pr50Gag protein expression; Gag protein purification; His-tag fusion protein
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This is an open access article distributed under the Creative Commons Attribution License which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited (CC BY 4.0).
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Krishnan, A.; Pillai, V.N.; Chameettachal, A.; Mohamed Ali, L.; Nuzra Nagoor Pitchai, F.; Tariq, S.; Mustafa, F.; Marquet, R.; A. Rizvi, T. Purification and Functional Characterization of a Biologically Active Full-Length Feline Immunodeficiency Virus (FIV) Pr50Gag. Viruses 2019, 11, 689.

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