Serum albumin binding molecules
The present invention relates to an antibody-like protein based on the tenth fibronectin type III domain ( 10 Fn3) that binds to serum albumin. The invention further relates to fusion molecules comprising a serum albumin-binding 10 Fn3 joined to a heterologous protein for use in diagnostic and therapeutic applications.
1. A method of producing a polypeptide comprising a fibronectin type III tenth ( 10 Fn3) domain that binds to domain 1 or 2 of human serum albumin with a K D of 1 μM or less, the method comprising:
(a) providing a host cell comprising a nucleic acid encoding the polypeptide, wherein the 10 Fn3 domain comprises a modified amino acid sequence in one or more of the BC, DE and FG loops relative to the wild-type human 10 Fn3 domain (SEQ ID NO: 1), and wherein the polypeptide comprises an amino acid sequence at least 75% identical to an amino acid sequence selected from the group consisting of SEQ ID NOs: 4, 8, 12, 16, 20, and 24-44, and
(b) culturing the host cell under conditions that allow expression of the polypeptide.
2. The method of claim 1 , further comprising isolating the polypeptide from the host cell.
3. The method of claim 1 , wherein the nucleic acid comprises a sequence selected from the group consisting of SEQ ID NOs: 176-214 and 401-405.
4. The method of claim 1 , wherein the nucleic acid is located within an expression vector.
5. The method of claim 1 , wherein the polypeptide comprises an amino acid sequence at least 80% identical to an amino acid sequence selected from the group consisting of: SEQ ID NOs: 4, 8,12, 16, 20, and 24-44.
6. The method of claim 5 , wherein the polypeptide comprises an amino acid sequence at least 85% identical to an amino acid sequence selected from the group consisting of: SEQ ID NOs: 4, 8, 12, 16, 20, and 24-44.
7. The method of claim 6 , wherein the polypeptide comprises an amino acid sequence at least 90% identical to an amino acid sequence selected from the group consisting of: SEQ ID NOs: 4, 8, 12, 16, 20, and 24-44.
8. The method of claim 7 , wherein the polypeptide comprises an amino acid sequence at least 95% identical to an amino acid sequence selected from the group consisting of: SEQ ID NOs: 4, 8,12, 16, 20, and 24-44.
9. The method of claim 8 , wherein the polypeptide comprises the amino acid sequence selected from the group consisting of SEQ ID NOs: 4, 8, 12, 16, 20, and 24-44.
10. The method of claim 1 , wherein the polypeptide comprises an amino acid sequence at least 75% identical to the amino acid sequence of SEQ ID NO: 4.
11. The method of claim 10 , wherein the polypeptide comprises an amino acid sequence at least 80% identical to the amino acid sequence of SEQ ID NO: 4.
12. The method of claim 11 , wherein the polypeptide comprises an amino acid sequence at least 85% identical to the amino acid sequence of SEQ ID NO: 4.
13. The method of claim 12 , wherein the polypeptide comprises an amino acid sequence at least 90% identical to the amino acid sequence of SEQ ID NO: 4.
14. The method of claim 13 , wherein the polypeptide comprises an amino acid sequence at least 95% identical to the amino acid sequence of SEQ 1D NO: 4.
15. The method of claim 14 , wherein the polypeptide comprises the amino acid sequence of SEQ ID NO: 4.
16. The method of claim 1 , wherein the polypeptide further comprises a heterologous protein.