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US 10,444,245 B2Granted
Trapping mammalian protein-protein complexes in virus-like particles utilizing HIV-1 GAG-bait fusion proteins
AssigneesVib Vzw, Universiteit Gent
InventorsJan Tavernier, Sven Eyckerman
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Abstract
The disclosure relates to a virus-like particle in which a protein complex is entrapped, ensuring the formation of the protein complex under physiological conditions, while protecting the protein complex during purification and identification. The disclosure further relates to the use of such virus-like particle for the isolation and identification of protein complexes.
Claims11
A method of detecting a protein-protein interaction in a cell, the method comprising:
- expressing in the cell a recombinant fusion protein comprising an HIV p55 GAG protein and a bait polypeptide of interest, wherein the expressed fusion protein forms a protein-protein complex with an endogenously-expressed prey polypeptide;
- forming virus-like particles comprising the protein-protein complex;
- isolating virus-like particles from the cell;
- and
- detecting specific bait-prey protein-protein binding in the virus-like particles as compared to background protein-protein interactions from a reference set of virus-like particles comprising HIV p55 GAG proteins not linked to the bait polypeptide of interest;
- thereby detecting the protein-protein interaction in the cell.
3 more elements
A method of detecting a protein-protein interaction in a cell, the method comprising:
- expressing in the cell recombinant fusion proteins comprising an HIV p55 GAG protein and a bait polypeptide of interest, wherein the expressed fusion proteins form a plurality of different protein-protein complexes with a plurality of different endogenously-expressed prey polypeptides;
- forming virus-like particles comprising the protein-protein complexes;
- isolating virus-like particles from the cell;
- and
- detecting specific bait-prey protein-protein binding in the virus-like particles as compared to background protein-protein interactions from a reference set of virus-like particles comprising HIV p55 GAG proteins not linked to the bait polypeptide of interest;
- thereby detecting the protein-protein interactions in the cell.
3 more elements
Dependencies are inferred from the claim text.