Purification and functional characterization of a biologically active full-length feline immunodeficiency virus (FIV) pr50gag

Anjana Krishnan, Vineeta N. Pillai, Akhil Chameettachal, Lizna Mohamed Ali, Fathima Nuzra Nagoor Pitchai, Saeed Tariq, Farah Mustafa, Roland Marquet, Tahir A. Rizvi

Research output: Contribution to journalArticlepeer-review

4 Citations (Scopus)

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.

Original languageEnglish
Article number689
JournalViruses
Volume11
Issue number8
DOIs
Publication statusPublished - Aug 2019

Keywords

  • Feline immunodeficiency virus (FIV)
  • Gag protein purification
  • His-tag fusion protein
  • Pr50 protein expression
  • Retroviral RNA packaging
  • Viral assembly

ASJC Scopus subject areas

  • Infectious Diseases
  • Virology

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