IP Library Granted Patent US 11,779,860
Granted Patent B2
US 11,779,860 · App. 16/637,323 · Granted Oct 10, 2023

Fc binding proteins with cysteine in the C-terminal helical region

Inventors: Erik Fiedler (Halle/Saale, DE); Ulrich Haupts (Halle/Saale, DE)
Assignee: Repligen Corporation
B01D15/3809B01J20/3274C07K1/22C07K14/31
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Quick Facts
Patent No.
US 11,779,860
App. No.
16/637,323
Granted
Oct 10, 2023
Kind
B2
Abstract

The present invention relates to Fc binding proteins comprising one or more domains with Cysteine in the C-terminal helical region. The invention further relates to affinity matrices comprising the Fc binding proteins of the invention. The invention also relates to a use of these Fc binding proteins or affinity matrices for affinity purification of immunoglobulins and to methods of affinity purification using the Fc binding proteins of the invention.

Claims (22)

1. An Fc binding protein comprising one or more domains, wherein the one or more domains comprises an amino acid sequence of any one of SEQ ID NOS: 3-86, 90-99 or an amino acid sequence with at least 89.5% identity to one of said sequences and wherein at least one amino acid in position 40, 42, 43, 46, 47, 49, 50, 51, 53, or 54 corresponding to SEQ ID NO: 3 is cysteine.

2. The Fc binding protein of claim 1 , wherein at least one domain comprises an amino acid sequence of SEQ ID NO: 3 or an amino acid sequence with at least 89.5% identity thereto wherein at least one amino acid in position 40, 42, 43, 46, 47, 49, 50, 51, 53, or 54 corresponding to SEQ ID NO: 3 is cysteine.

3. The Fc binding protein of claim 1 , wherein at least one domain comprises an amino acid sequence of SEQ ID NOs: 3-16 or an amino acid sequence with at least 89.5% identity thereto, respectively, wherein at least one amino acid in position 40, 42, 43, 46, 47, 50, 51, 53, or 54 corresponding to SEQ ID NOs: 3-16 is cysteine.

4. The Fc binding protein of claim 1 , wherein at least one domain comprises an amino acid sequence of SEQ ID NOs: 7-8 or an amino acid sequence with at least 89.5% identity thereto, respectively, wherein at least one amino acid in position 40, 42, 43, 46, 47, 50, 51, 53, or 54 corresponding to SEQ ID NOs: 7-8 is cysteine.

5. The Fc binding protein of claim 1 , wherein at least one amino acid in position 43, 46, or 47 corresponding to SEQ ID NO: 3 is cysteine.

6. The Fc binding protein of claim 1 , wherein at least one domain comprises an amino acid sequence selected from the group consisting of SEQ ID NOs: 17-86, 90-99, or an amino acid sequence with at least 89.5% identity thereto, respectively.

7. The Fc binding protein of claim 1 , wherein said domain has a deletion of 1, 2, or 3 amino acids within the first 4 amino acids of its N-terminus and/or a deletion of 1 or 2 amino acids within the first 2 amino acids of its C-terminus.

8. The Fc binding protein of claim 1 , wherein the protein comprises 2, 3, 4, 5, 6, 7, or 8 domains linked to each other.

9. The Fc binding protein of claim 8 , wherein the protein is a homo-multimer or a hetero-multimer.

10. The Fc binding protein of claim 9 , wherein one or more domains are linked to each other directly or with one or more linkers.

11. The Fc binding protein of claim 1 , wherein the protein is conjugated to a solid support.

12. The Fc binding protein of claim 1 , wherein said protein binds to IgG1, IgG2, IgG4, IgM, IgA, Ig fragments comprising an Fc region, fusion proteins comprising an Fc region of an Ig, and conjugates comprising an Fc region of an Ig.

13. An affinity separation matrix comprising an Fc binding protein comprising one or more domains, wherein the one or more domains comprises an amino acid sequence of any one of SEQ ID NOS: 3-16 or an amino acid sequence with at least 89.5% identity to one of said sequences and wherein at least one amino acid in position 40, 42, 43, 46, 47, 49, 50, 51, 53, or 54 corresponding to SEQ ID NO: 3 is cysteine.

14. The affinity separation matrix of claim 13 , wherein at least one domain comprises an amino acid sequence of SEQ ID NO: 3 or an amino acid sequence with at least 89.5% identity thereto wherein at least one amino acid in position 40, 42, 43, 46, 47, 49, 50, 51, 53, or 54 corresponding to SEQ ID NO: 3 is cysteine.

15. The affinity separation matrix of claim 13 , wherein at least one domain comprises an amino acid sequence of SEQ ID NOs: 7-8 or an amino acid sequence with at least 89.5% identity thereto, respectively, wherein at least one amino acid in position 40, 42, 43, 46, 47, 50, 51, 53, or 54 corresponding to SEQ ID NOs: 7-8 is cysteine.

16. The affinity separation matrix of claim 13 , wherein at least one amino acid in position 43, 46, or 47 corresponding to SEQ ID NO: 3 is cysteine.

17. A method of affinity purification of a protein comprising an Fc sequence, the method comprising: (a) providing a liquid that contains protein comprising an Fc sequence; (b) providing an affinity separation matrix comprising an Fc binding protein comprising one or more domains, wherein the one or more domains comprises an amino acid sequence of any one of SEQ ID NOS: 3-16 or an amino acid sequence with at least 89.5% identity to one of said sequences and wherein at least one amino acid in position 40, 42, 43, 46, 47, 49, 50, 51, 53, or 54 corresponding to SEQ ID NO: 3 is cysteine, wherein the Fc binding protein is coupled to said affinity separation matrix; (c) contacting said affinity separation matrix with the liquid under conditions that permit binding of the Fc binding protein to a protein comprising an Fc sequence; and (d) eluting said protein comprising an Fc sequence from said affinity separation matrix.

18. The method of claim 17 , wherein at least one domain comprises an amino acid sequence of SEQ ID NO: 3 or an amino acid sequence with at least 89.5% identity thereto wherein at least one amino acid in position 40, 42, 43, 46, 47, 49, 50, 51, 53, or 54 corresponding to SEQ ID NO: 3 is cysteine.

19. The method of claim 17 , wherein at least one domain comprises an amino acid sequence of SEQ ID NOs: 7-8 or an amino acid sequence with at least 89.5% identity thereto, respectively, wherein at least one amino acid in position 40, 42, 43, 46, 47, 50, 51, 53, or 54 corresponding to SEQ ID NOs: 7-8 is cysteine.

20. The method of claim 17 , wherein at least one amino acid in position 43, 46, or 47 corresponding to SEQ ID NO: 3 is cysteine.

21. The Fc binding protein of claim 10 , wherein said one or more linkers is a peptide linker.

22. The Fc binding protein of claim 11 , wherein the protein is conjugated to a solid support via a cysteine in position 40, 42, 43, 46, 47, 50, 51, 53, or 54.

Assignments (1)
ASSIGNMENT OF ASSIGNOR'S INTEREST Recorded May 7, 2020
From: FIEDLER, ERIK; HAUPTS, ULRICH
To: REPLIGEN CORPORATION
Reel/Frame 052597/0194 →
Priority Claims (3)
WO PCT/EP2017/069976 · Aug 7, 2017 · international
EP 18154731 · Feb 1, 2018 · regional
EP 18163964 · Mar 26, 2018 · regional
Continuity (1)
Related Publication 20200238197A1 · Jul 30, 2020
Cited By (1)
US 12,577,276