Hepatitis c virus E1/E2 heterodimers and methods of producing same
AssigneeThe Governors of the University of Alberta
InventorsMichael Houghton, Darren Hockman, John L. Law, Chao Chen, Michael Logan
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Abstract
The present disclosure provides affinity tagged heterodimeric polypeptides comprising a hepatitis C virus (HCV) E1 polypeptide and an HCV E2 polypeptide, where one or both of the E1 and E2 polypeptides comprises an affinity tag. The present disclosure provides a method of producing an affinity tagged E1/E2 heterodimer of the present disclosure. The present disclosure provides methods of producing untagged HCV E1/E2 heterodimers. The present disclosure provides HCV E1/E2 heterodimers, compositions comprising same, and methods of inducing an immune response to HCV.
Claims15
A method of purifying a hepatitis C virus (HCV) E1/E2 heterodimer from a lysate, the method comprising:
- a) contacting a lysate of a genetically modified host cell with an affinity tag-binding polypeptide immobilized on an insoluble support, wherein the genetically modified host cell is genetically modified with a nucleic acid comprising a nucleotide sequence encoding a polyprotein comprising, in order from N-terminus to C-terminus:
- a) an HCV E1 polypeptide;
- and
- b) an HCV E2-affinity tag fusion polypeptide comprising, in order from N-terminus to C-terminus:
- i) an affinity tag polypeptide;
- ii) a proteolytically cleavable linker;
- and iii) an HCV E2 polypeptide
- i) a first nucleotide sequence encoding an HCV E1 polypeptide;
- and
- ii) a second nucleotide sequence encoding an HCV E2 polypeptide,
- wherein the polyprotein is processed in the host cell to produce the affinity tagged HCV E1/E2 heterodimer and the lysate comprises the affinity tagged HCV E1/E2 heterodimer,
- wherein the affinity tagged HCV E1/E2 heterodimer present in the lysate binds to the immobilized affinity tag-binding polypeptide, generating an immobilized affinity tagged HCV E1/E2 heterodimer;
- b) contacting the immobilized HCV E1/E2 heterodimer with a protease that cleaves the proteolytically cleavable linker present in the immobilized affinity tagged HCV E1/E2 heterodimer, thereby releasing the HCV E1/E2 heterodimer from the affinity tag;
- and
- c) collecting the HCV E1/E2 heterodimer.
Dependencies are inferred from the claim text.