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Purification and Functional Characterization of a Biologically Active Full-Length Feline Immunodeficiency Virus (FIV) Pr50Gag
- Source :
- Viruses, Volume 11, Issue 8, Viruses, MDPI, 2019, 11 (8), pp.689. ⟨10.3390/v11080689⟩, Viruses, Vol 11, Iss 8, p 689 (2019)
- Publication Year :
- 2019
- Publisher :
- Multidisciplinary Digital Publishing Institute, 2019.
-
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.
- Subjects :
- 0301 basic medicine
retroviral RNA packaging
Feline immunodeficiency virus
[SDV]Life Sciences [q-bio]
viruses
lcsh:QR1-502
Gene Expression
Gene Products, gag
Genome, Viral
Immunodeficiency Virus, Feline
Pr50Gag protein expression
Virus
lcsh:Microbiology
Article
His-tag fusion protein
law.invention
viral assembly
03 medical and health sciences
law
Virology
feline immunodeficiency virus (FIV)
Escherichia coli
Animals
Humans
[SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology
Guide RNA
030102 biochemistry & molecular biology
biology
Chemistry
Virus Assembly
RNA
Biological activity
[SDV.BBM.BM]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular biology
biology.organism_classification
Fusion protein
In vitro
Recombinant Proteins
030104 developmental biology
Infectious Diseases
HEK293 Cells
Biochemistry
Recombinant DNA
Cats
RNA, Viral
Pr50 Gag protein expression
Gag protein purification
Subjects
Details
- Language :
- English
- ISSN :
- 19994915
- Database :
- OpenAIRE
- Journal :
- Viruses
- Accession number :
- edsair.doi.dedup.....dea5b10227dea11314c4aa5a10c79765
- Full Text :
- https://doi.org/10.3390/v11080689