Circular tandem repeat proteins
Circular handed alpha-helical repeat proteins are described. The repeat proteins have a number of uses as scaffolds for geometrically precise, arrayed presentation of cell-signaling or immune-related protein and peptide epitopes, as well as numerous other therapeutic, diagnostic, and nanotechnological uses.
1. A protein having the formula: (a-b-x-y) n wherein
a and x represent linker sequences;
b represents an amino acid sequence that forms an alpha (α) helix;
y represents an amino acid sequence that forms a second α helix;
n=3, 6, 9, 12, or 24;
the protein is handed, and
the N- and C-termini of the protein create a circular architecture.
2. The protein of claim 1 , wherein the linker sequences are flexible linker sequences.
3. The protein of claim 2 , wherein the flexible linker sequences are GBB linker sequences.
4. The protein of claim 3 , wherein the GBB linker sequences are selected from GKS; GIT; GTT; GYS; GDK; GDE; NDK; GDR; GDL; and GIS.
5. The protein of claim 1 , wherein the protein is left-handed.
6. The protein of claim 1 , wherein b and y have 100% sequence identity.
7. The protein of claim 1 , wherein b and y have at least 98% sequence identity.
8. The protein of claim 1 , wherein b and y are selected from the sequences as set forth in SEQ ID NOs. 1-50; 124-129; 139; 140; 146; or 147.
9. A protein of claim 1 , wherein each (a-b-x-y) unit that is not an N-terminal or C-terminal (a-b-x-y) unit has 100% sequence identity with an adjacent (a-b-x-y) unit.
10. A protein of claim 1 , wherein each (a-b-x-y) unit has at least 95% sequence identity with an adjacent (a-b-x-y) unit.
11. The protein of claim 1 , comprising a sequence selected from the sequences as set forth in SEQ ID NOs. 51-70; 117-123; 135-138 or 145.
12. A circular, handed protein comprising at least three α-helical structures wherein each α-helical structure comprises an outer α helix and an inner α helix joined by a flexible linker and wherein each α-helical structure has at least 95% sequence identity with an adjacent α-helical structure.
13. The circular, handed protein of claim 12 , wherein the flexible linker is a GBB linker.
14. The circular, handed protein of claim 13 , wherein the GBB linker is a sequence selected from GKS; GIT; GTT; GYS; GDK; GDE; NDK; GDR; GDL; and GIS.
15. The circular, handed protein of claim 12 , further comprising at least two functional domains wherein each functional domain is inserted into the sequence of the protein between an outer α helix of the protein and an adjacent inner α helix of the protein and wherein at least one of the functional domains is selected from a cytokine, a Notch ligand, an immunogenic peptide, a peptide adjuvant, a single-chain class I MHC domain, or a small molecule ligand binding domain.
16. A protein having the formula: (d-a-b-x-y) n ; (a-d-b-x-y) n ; (a-b-d-x-y) n ; (a-b-x-d-y) n or (a-b-x-y-d) n wherein
a and x represent linker sequences;
b represents an amino acid sequence that forms an alpha (α) helix;
y represents an amino acid sequence that forms a second α helix;
d represents a functional domain;
n=3, 6, 9, 12, or 24;
the protein is handed, and
the N- and C-termini of the protein create a circular architecture.
17. The protein of claim 16 , wherein the functional domain comprises a cytokine, a Notch ligand, an immunogenic peptide, a peptide adjuvant, a single-chain class I MHC domain, or a small molecule ligand binding domain.
18. The protein of claim 16 , wherein the functional domain comprises SH2, SH3, IL-2, IL-3, IL-17c, single-chain MHC, the extracellular domain of the Delta-1 Notch protein ligand, Protein L, a protein having the sequence set forth in SEQ ID NO: 116, or a protein having the sequence set forth in SEQ ID NO: 115.
19. The protein of claim 16 , wherein the linker sequences are flexible linker sequences.
20. The protein of claim 16 , wherein b and y have at least 98% sequence identity.