Modified lipopolysaccharide glycoform and method of use

Patent metadata

Field	Value
Publication number	US-10864266-B2
Application number	US-201916690739-A
Country	US
Kind code	B2
Filing date	Nov 21, 2019
Priority date	May 9, 2014
Publication date	Dec 15, 2020
Grant date	Dec 15, 2020

Abstract

The present disclosure generally relates to genetic engineering of bacteria. More particularly, the present disclosure describes genetic engineering of E. coli to create mutant O-antigen ligase, as well as novel lipopolysaccharide molecules resulting from that genetic engineering. Methods for using those novel molecules are also described.

First claim

What is claimed is:

1. A vaccine adjuvant comprising at least one of a lipopolysaccharide isolated from a mutant O-antigen ligase or lipopolysaccharide derivative isolated from the mutant O-antigen ligase, the mutant O-antigen ligase comprising either an isolated protein that has the amino acid sequence SEQ ID NO: 1, or
an isolated protein having at least 90% sequence identity to SEQ ID NO: 2 and having an amino acid substitution of phenylalanine to serine at the phenylalanine homologous to position 332 of SEQ ID NO: 2,
wherein the isolated protein has at least one of 0-antigen ligase activity or the activity to ligate lipid A to disaccharide pentapeptide (DPP).

2. A vaccine adjuvant comprising a lipopolysaccharide derivative molecule prepared by the process comprising the steps of:
providing an E. coli strain adapted to express an LPS derivative molecule;
modifying the E. coli strain by creating a mutant 0-antigen ligase selected from the group consisting of:
a mutant O-antigen ligase comprising an isolated protein that has the amino acid sequence SEQ ID NO: 1, and
a mutant O-antigen ligase comprising an isolated protein having at least 90% sequence identity to SEQ ID NO: 2 and having an amino acid substitution of phenylalanine to serine at the phenylalanine homologous to position 332 of SEQ ID NO: 2, wherein the protein has at least one of 0-antigen ligase activity or the activity to ligate lipid A to disaccharide pentapeptide (DPP);

providing a growth media for the modified E. coli strain; and
allowing the modified E. coli strain to produce an LPS* derivative molecule, wherein the LPS* derivative molecule includes at least one non-native sugar not present in an LPS molecule it derives from.

Assignees

• Univ Princeton
• Harvard College

Inventors

• Silhavy Thomas J
• Grabowicz Marcin
• Kahne Daniel
• Lebar Matthew
• Andres Dorothee

Classifications

• C07H13/06

  Fatty acids · CPC title

• C12N15/04

  Fungi · CPC title

• A61K2039/55572

  Lipopolysaccharides; Lipid A; Monophosphoryl lipid A · CPC title

• C12N9/00

  Enzymes; Proenzymes; Compositions thereof (preparations containing enzymes for cleaning teeth A61K8/66, A61Q11/00; medicinal preparations containing enzymes or proenzymes A61K38/43; enzyme containing detergent compositions C11D; {enzymes with nucleic acid structure, e.g. ribozymes, C12N15/113}); Processes for preparing, activating, inhibiting, separating or purifying enzymes (preparation of malt C12C1/00) · CPC title

• A61K2039/55516

  Proteins; Peptides · CPC title