Mutated immunoglobulin-binding polypeptides

Patent metadata

Field	Value
Publication number	US-9896486-B2
Application number	US-201414903837-A
Country	US
Kind code	B2
Filing date	Jul 8, 2014
Priority date	Jul 10, 2013
Publication date	Feb 20, 2018
Grant date	Feb 20, 2018

Abstract

A polypeptide with improved alkaline stability, which polypeptide comprises a mutant of a B or C domain of Staphylococcus Protein A, as specified by SEQ ID NO 1 or SEQ ID NO 2, or of Protein Z, as specified by SEQ ID NO 3, wherein at least the glutamine residue at position 9 has been mutated to an amino acid other than asparagine. The invention also discloses multimers of said polypeptide, as well as separation matrices comprising the multimers or polypeptides.

First claim

What is claimed is:

1. A polypeptide with improved alkaline stability, which polypeptide comprises a mutant of a B or C domain of Staphylococcus Protein A (SpA), as specified by SEQ ID NO 1 or SEQ ID NO 2, or of Protein Z, as specified by SEQ ID NO 3, wherein at least the glutamine residue at position 9 has been mutated to an amino acid other than asparagine, comprising a sequence selected from the group consisting of: SEQ ID NO 6, SEQ ID NO 7, SEQ ID NO 8, SEQ ID NO 9, SEQ ID NO 10, SEQ ID NO 20, SEQ ID NO 25, SEQ ID NO 26 and SEQ ID NO 27.

2. The polypeptide or multimer according claim 1 , further comprising at the C-terminal or N-terminal one or more coupling element, selected from the group consisting of a cysteine residue, a plurality of lysine residues and a plurality of histidine residues.

3. A nucleic acid or a vector encoding a polypeptide or multimer according to claim 1 .

4. A separation matrix, wherein a plurality of polypeptides or multimers according to claim 1 have been coupled to a solid support.

5. A method of isolating an immunoglobulin, wherein a separation matrix according to claim 4 is used.

6. The method of claim 5 , comprising the steps of:
contacting a liquid sample comprising an immunoglobulin with the separation matrix,
washing said separation matrix with a washing liquid,
eluting the immunoglobulin from the separation matrix with an elution liquid, and cleaning the separation matrix with a cleaning liquid.

7. The polypeptide of claim 1 , wherein the sequence is SEQ ID NO 6.

8. The polypeptide of claim 1 , wherein the sequence is SEQ ID NO 7.

9. The polypeptide of claim 1 , wherein the sequence is SEQ ID NO 8.

10. The polypeptide of claim 1 , wherein the sequence is SEQ ID NO 9.

11. The polypeptide of claim 1 , wherein the sequence is SEQ ID NO 10.

12. The polypeptide of claim 1 , wherein the sequence is SEQ ID NO 20.

13. The polypeptide of claim 1 , wherein the sequence is SEQ ID NO 25.

14. The polypeptide of claim 1 , wherein the sequence is SEQ ID NO 26.

15. The polypeptide of claim 1 , wherein the sequence is SEQ ID NO 27.

Assignees

• Ge Healthcare Bioprocess R&D Ab

Inventors

• Rodrigo Gustav
• Ander Mats
• Bjorkman Tomas
• Bauren Goran

Classifications

• B01J20/3212

  consisting of a polymer obtained by reactions otherwise than involving only carbon to carbon unsaturated bonds · CPC title

• C07K1/22

  Affinity chromatography or related techniques based upon selective absorption processes · CPC title

• B01J20/3219

  involving a particular spacer or linking group, e.g. for attaching an active group · CPC title

• C07K14/31 Primary

  from Staphylococcus (G) · CPC title

• B01D15/3809

  of the antigen-antibody type, e.g. protein A, G or L chromatography · CPC title