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 comprising the formula:
[(d-a-b-x-y) 1 (a-b-x-y) 5 ] 4
wherein d is a single-chain protein domain comprising a class I major histocompatibility complex (MHC)-peptide complex;
a is GLN;
x is GIS;
b is SEQ ID NO: 124;
y is SEQ ID NO: 125 or SEQ ID NO: 129;
the protein is left-handed; and
the N- and C-termini of the protein create a circular architecture.
2. The protein of claim 1 , wherein each (a-b-x-y) unit has the sequence set forth in SEQ ID NO: 130 or SEQ ID NO: 134.
3. The protein of claim 1 , wherein each (a-b-x-y) unit that is not an end unit of the protein has 100% sequence identity with an adjacent (a-b-x-y) unit.
4. The protein of claim 1 , wherein the protein exhibits high thermostability, high solubility, and is self-folding.
5. The protein of claim 1 , wherein the N- and C-termini of the protein create a closed architecture.
6. A protein having the formula: (a-b-x-y)n wherein
a represents a linker sequence;
x represents a linker sequence;
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=2 or more;
each (a-b-x-y) unit is structurally repetitive to an adjacent (a-b-x-y) unit,
the protein is left-handed;
the N- and C-termini of the protein create a circular architecture, and
wherein the feature of structurally repetitive is determined by stacking each (a-b-x-y) unit with an adjacent (a-b-x-y) unit, and after stacking, the distance between corresponding atoms of the stacked outer α-helices and the stacked inner α-helices is within 2 angstrom (Å) measured using root-mean-square deviation (RMSD).
7. The protein of claim 6 , wherein b has the sequence set forth in SEQ ID NO: 124 in at least one (a-b-x-y) unit.
8. The protein of claim 6 , wherein y has the sequence set forth in SEQ ID NO: 125 in at least one (a-b-x-y) unit.
9. The protein of claim 6 , wherein y has the sequence set forth in SEQ ID NO: 129 in at least one (a-b-x-y) unit.
10. The protein of claim 6 , wherein b and y are selected from the sequence set forth in SEQ ID NOs. 1-50; 124-129; 139; 140; 146; or 147 provided that b and y are not the same sequence.
11. The protein of claim 6 , wherein at least one (a-b-x-y) unit has the sequence set forth in SEQ ID NO: 130.
12. The protein of claim 6 , wherein at least one (a-b-x-y) unit has the sequence set forth in SEQ ID NO: 134.
13. The protein of claim 6 , wherein a (a-b-x-y) unit has the sequence set forth in Previously Presented SEQ ID NOs. 73-114; 130-134; 141-144; 172; or 148.
14. The protein of claim 6 , wherein the linker sequences comprise GBB linker sequences.
15. The protein of claim 14 , wherein the GBB linker sequences are selected from GKS; GIT; GTT; GYS; GDK; GDE; NDK; GDR; GDL; and GIS.
16. The protein of claim 6 , comprising a functional domain (d) at the N-terminus of at least one (a-b-x-y) unit.
17. The protein of 68, wherein the functional domain comprises SH2, SH3, IL-2, IL-3, IL-17c, a single-chain MHC domain, the extracellular domain of the Delta-1 Notch protein ligand, Protein L, the sequence set forth in SEQ ID NO: 116, or the sequence set forth in SEQ ID NO: 115.
18. The protein of claim 6 , wherein n=24.
19. The protein of claim 18 , comprising a functional domain (d) at the N-terminus of the first (a-b-x-y) unit, the N-terminus of the seventh (a-b-x-y) unit, the N-terminus of the thirteenth (a-b-x-y) unit, and the N-terminus of the nineteenth (a-b-x-y) unit.
20. The protein of 71, wherein the functional domain comprises SH2, SH3, IL-2, IL-3, IL-17c, a single-chain MHC domain, the extracellular domain of the Delta-1 Notch protein ligand, Protein L, or the sequence set forth in SEQ ID NO: 116, or the sequence set forth in SEQ ID NO: 115.