Methods of host cell modification

Patent metadata

Field	Value
Publication number	US-12398380-B2
Application number	US-202117541454-A
Country	US
Kind code	B2
Filing date	Dec 3, 2021
Priority date	Oct 11, 2013
Publication date	Aug 26, 2025
Grant date	Aug 26, 2025

Abstract

Described herein are methods of inserting nucleic acid sequences into host cells. Also described herein are genetically stable host cells comprising inserted nucleic acid sequences and methods of using such host cells in the generation of proteins.

First claim

What is claimed:

1. A method of producing an N-glycosylated protein, wherein the method comprises culturing an Escherichia coli ( E. coli ) host cell under conditions suitable for production of proteins, wherein the E. coli host cell genome comprises:
(i) a heterologous DNA sequence that encodes an oligosaccharyl transferase;
(ii) a heterologous DNA sequence of at least 12 kb that encodes a glycosyltransferase from Candida albicans wherein the glycosyltransferase from Candida albicans is capable of making a Candida albicans polysaccharide; and
wherein the E. coli host cell further comprises a gene that encodes a carrier protein containing at least SEQ ID NO: 72 having the sequence D/E-X-N-Z-S/T, wherein X and Z are independently selected from any amino acid except proline.

2. The method of claim 1 , wherein the carrier protein is detoxified Exoprotein A from Pseudomonas aeruginosa (EPA).

3. The method of claim 1 , wherein the heterologous DNA sequence that encodes an oligosaccharyl transferase comprises a pglB gene.

4. The method of claim 3 , wherein the pglB gene is from C. jejuni.

5. The method of claim 1 , wherein the glycosyltransferase from Candida albicans is capable of making a polysaccharide from Candida albicans.

Assignees

• Glaxosmithkline Biologicals Sa
• Glaxosmithkline Biologicals Sa

Inventors

• Wacker Michael
• Kowarik Michael
• Fernandez Fabiana

Classifications

• C12P21/00

  Preparation of peptides or proteins (single cell protein C12N1/00) · CPC title

• Y02A50/30

  Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change · CPC title

• A61K39/0258

  Escherichia · CPC title

• C12N2800/30

  Vector systems comprising sequences for excision in presence of a recombinase, e.g. loxP or FRT · CPC title

• C12N2795/00041

  Use of virus, viral particle or viral elements as a vector · CPC title