Covalently fused viral coat proteins for the display of target molecules
Inventors
Tottey, Stephen • Musiychuk, Konstantin
Assignees
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Publication Number
US-11618771-B2
Publication Date
2023-04-04
Expiration Date
Abstract
A fusion protein comprising a target protein, a first recombinant viral coat protein, a second recombinant viral coat protein and a first linkage peptide is provided. The target protein is at N-terminus of the first recombinant viral coat protein. The first recombinant viral coat protein is linked to N-terminus of the first linkage peptide. The second recombinant viral coat protein is linked to C-terminus of the first linkage peptide. The first and second recombinant viral coat proteins are derived from the coat protein (CP) of alfalfa mosaic virus (AIMV). The fusion protein may further comprise a second linkage peptide between the target protein and the first recombinant viral coat protein. The fusion protein may form a virus like particle (VLP). The target protein may be displayed on the surface of the VLP. Also provided are methods for producing the fusion protein and the VLP as well as the uses of the fusion protein and/or the VLP.
Core Innovation
The invention provides a fusion protein that comprises a target protein, a first recombinant viral coat protein, a second recombinant viral coat protein, and a first linkage peptide. The target protein is positioned at the N-terminus of the first recombinant viral coat protein, the first recombinant viral coat protein is linked to the N-terminus of the first linkage peptide, and the second recombinant viral coat protein is linked to the C-terminus of the first linkage peptide.
The recombinant viral coat proteins are based on an AIMV coat protein sequence and comprise an amino acid sequence identical to SEQ ID NO: 1 or at least 80% identical to SEQ ID NO: 1 with one or more deletions, insertions or substitutions within residues 17-38 of SEQ ID NO: 1. The arrangement supports defined placement of both coat proteins relative to the linkage peptide and target protein.
The disclosure further describes virus-like particles based on the fusion protein, including embodiments where the target protein is displayed on the surface of the virus-like particle. The disclosed constructs are used as immunological, therapeutic, diagnostic, and enzyme-catalyzed agents, and include examples such as Pfs25, Pfs230, influenza hemagglutinin subdomains, Western Equine encephalitis virus antigen, Eastern Equine encephalitis virus antigen, and horseradish peroxidase.
Claims Coverage
The independent claim defines the structural fusion protein with two recombinant viral coat proteins linked through a linkage peptide around an N-terminally positioned target protein. The claim includes sequence-constraining branches that specify identity and allowable edits within residues 17-38 of SEQ ID NO: 1.
N-terminally positioned target between first and second recombinant viral coat proteins via a linkage peptide
A fusion protein comprising a target protein, a first recombinant viral coat protein, a second recombinant viral coat protein and a first linkage peptide, wherein the target protein is at N-terminus of the first recombinant viral coat protein, wherein the first recombinant viral coat protein is linked to N-terminus of the first linkage peptide, and wherein the second recombinant viral coat protein is linked to C-terminus of the first linkage peptide.
First recombinant viral coat protein constrained to SEQ ID NO: 1 identity with edits in residues 17-38
The first recombinant viral coat protein comprises an amino acid sequence identical to SEQ ID NO: 1 or at least 80% identical to SEQ ID NO: 1 with one or more deletions, insertions or substitutions within residues 17-38 of SEQ ID NO: 1.
Second recombinant viral coat protein constrained to SEQ ID NO: 1 identity with edits in residues 17-38
The second recombinant viral coat protein comprises an amino acid sequence identical to SEQ ID NO: 1 or at least 80% identical to SEQ ID NO: 1 with one or more deletions, insertions or substitutions within residues 17-38 of SEQ ID NO: 1.
The key inventive coverage is the specific N-terminal placement of the target protein and the linking topology that places the first recombinant viral coat protein at the N-terminus of the linkage peptide and the second recombinant viral coat protein at the C-terminus, combined with sequence constraints requiring each coat protein to be SEQ ID NO: 1 (or at least 80% identical) with defined allowable edits within residues 17-38.
Stated Advantages
Not explicitly described in patent.
Documented Applications
Immunological response induction, disease treatment, biomarker detection, and enzyme-catalyzed reactions using the disclosed fusion proteins and virus-like particles.
Diagnostic binding for biomarker detection by contacting a sample with an effective amount of the fusion protein or virus-like particles made from the fusion protein.
Enzyme-catalyzed reactions by contacting a reagent with an effective amount of a fusion protein or virus-like particles made from the fusion protein.
Example antigen/enzyme targets include malaria antigens Pfs25 and Pfs230, influenza hemagglutinin subdomains (H1/H3/H5/H7), Western Equine encephalitis virus antigen, Eastern Equine encephalitis virus antigen, and horseradish peroxidase (HRP).
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