Supramolecular filamentous assemblies for protein purification

Patent metadata

Field	Value
Publication number	US-12357965-B2
Application number	US-202318460024-A
Country	US
Kind code	B2
Filing date	Sep 1, 2023
Priority date	Aug 18, 2017
Publication date	Jul 15, 2025
Grant date	Jul 15, 2025

Abstract

The present invention provide novel immunofiber compositions for protein or peptide purification and simple and cost-efficient methods and systems using these compositions. In some embodiments, the immunofibers comprise a customized Z-33 peptide derived from Staphylococcus aureus Protein A which is used to construct immuno-amphiphile molecules that assemble into immunofibers in aqueous solution with bioactive epitopes on the surface and have peptide or protein binding ability.

First claim

The invention claimed is:

1. An immunofiber composition comprising:
a) a spacer molecule having a hydrocarbon chain at its N-terminus conjugated to a peptide sequence comprising the generic amino acid sequence of XXBB,
wherein XX is two amino acids having a small hydrophobic side chain and can be the same or different amino acid, and
wherein BB is two amino acids having a negatively charged side chain and can be the same or different amino acid; and

b) an immunofiber binding molecule comprising an antibody binding peptide conjugated to a second hydrocarbon chain,
wherein the antibody binding peptide has a hydrophilic amino acid sequence selected from the group consisting of SEQ ID NOS: 1-7.

2. The immunofiber composition of claim 1 , wherein each of the amino acids having a small hydrophobic side chain is independently selected from the group consisting of: Ala, Val, Ile, and Leu.

3. The immunofiber composition of claim 1 , wherein each of the amino acids having a negatively charged side chain is independently selected from the group consisting of: Glu and Asp.

4. The immunofiber composition of claim 1 , wherein XXBB is VVEE (SEQ ID NO: 10).

5. The immunofiber composition of claim 1 , wherein the hydrocarbon chain is a C12 hydrocarbon chain.

6. The immunofiber composition of claim 1 , wherein the antibody binding peptide has the amino acid sequence FNMQQQRRFYEALHDPNLNEEQRNAKIKSIRDD (SEQ ID NO: 1).

7. The immunofiber composition of claim 1 , wherein the second hydrocarbon chain is between 8 and 22 carbons in length and is either linear or branched.

8. The immunofiber composition of claim 1 , wherein the second hydrocarbon chain is between 8 and 12 carbons in length.

9. A method for purifying antibodies or Fc-containing peptides or proteins, said method comprising:
a) dissolving the immunofiber composition of claim 1 in an aqueous solution at a physiological pH to provide an immunofiber solution and aging the immunofiber solution overnight to provide a solution of self-assembled immunofibers;
b) mixing a sample comprising antibodies or Fc-containing peptides or proteins with the solution of self-assembled immunofibers to provide a solution comprising immunofiber/antibody complexes or immunofiber/Fc-containing peptide or protein complexes;
c) separating the immunofiber/antibody complexes or the immunofiber/Fc-containing peptide or protein complexes from the solution by adding salt and centrifuging; and
d) dissociating the immunofibers from the antibodies or Fc-containing peptide or protein complexes and collecting the unbound antibodies or Fc-containing peptides or proteins.

10. The method of claim 9 , wherein aging the immunofiber solution overnight promotes binding of the immunofibers to the Fc portions of the antibodies or Fc-containing peptides or proteins.

11. The method of claim 9 , wherein dissociating the immunofibers from the antibodies or Fc-containing peptides or proteins comprises lowering the pH to an elution condition; and using one or more of filtration, microfiltration, or ultrafiltration.

12. The method of claim 9 , further comprising further purifying the antibodies or Fc-containing peptides or proteins using polishing.

Assignees

• Univ Johns Hopkins
• Bristol Myers Squibb Co

Inventors

• Cui Honggang
• Li Yi
• Lock Lye Lin
• Xu Xuankuo
• Li Zhengjian

Classifications

• C07K7/08

  having 12 to 20 amino acids (gastrins C07K14/595; somatostatins C07K14/655; melanotropins C07K14/68) · CPC title

• C07K16/065

  Purification, fragmentation · CPC title

• C07K14/31 Primary

  from Staphylococcus (G) · CPC title

• C07K1/22

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

• B01J20/3475

  in the liquid phase · CPC title