Polysaccharide and uses thereof
AssigneeGlaxosmithkline Ciologicals S.A.
InventorsMichael T. Kowarik, Michael L. Wetter, Stefan J. Kemmler, Micha A. Häuptle, Veronica Gambillara, Manuela Mally
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Abstract
Provided herein is a novel E. coli O polysaccharide, O25B. Also provided herein are prokaryotic host cells containing enzymes (e.g., glycosyltransferases) used in O25B production. The host cells provided herein produce O25B bioconjugates, wherein said bioconjugates contain O25B linked to a carrier protein. Further provided herein are compositions, e.g., pharmaceutical compositions, including O25B and/or bioconjugates containing O25B. Such compositions can be used as vaccines against infection with ExPEC, and may further include one or more additional bioconjugates.
Claims20
A composition comprising a bioconjugate of an E. coli O25B antigen covalently coupled to a carrier protein, wherein the E. coli O25B antigen comprises the structure of Formula O25B′:
- wherein n is an integer of 1 to 30.
A recombinant prokaryotic host cell for producing a bioconjugate of an E. coli O25B antigen covalently coupled to a carrier protein, comprising:
- (a) a nucleotide sequence comprising an rfb gene cluster encoding the E. coli O25B antigen, wherein the rfb gene cluster encodes:
- i. dTDP-Glucose 4,6-dehydratase;
- ii. dTDP-6-Deoxy-D-glucose 3,5-epimerase;
- iii. Glucose-1-phosphate thymidylyltransferase;
- iv. dTDP-4-dehydrorhamnose 3,5-epimerase;
- v. O antigen flippase;
- vi. dTDP-Rha:Glc-Rha(Ac)-GlcNAc-UPP α-1,3-rhamnosyltransferase;
- vii. UDP-Glc:Glc-Rha(Ac)-GlcNAc-UPP β-1,6-glucosyltransferase;
- viii. O antigen polymerase;
- ix. O-acetyl transferase;
- x. UDP-Glc:Rha-GlcNAc-UPP α-1,3-glucosyltransferase;
- and
- xi. dTDP-Rha:
- GlcNAc-UPP α-1,3-rhamnosyltransferase;
- (b) a nucleotide sequence encoding an oligosaccharyl transferase;
- and
- (c) a nucleotide sequence encoding the carrier protein, wherein the carrier protein comprises a consensus sequence Asn-X-Ser(Thr), and X is any natural amino acid except Pro (SEQ ID NO:
- 14).
A bioconjugate of an E. coli O25B antigen covalently coupled to a carrier protein,
- wherein the bioconjugate is produced by a method comprising:
- (a) culturing a recombinant prokaryotic host cell under conditions for production of the bioconjugate, the recombinant prokaryotic host cell comprising:
- i. a nucleotide sequence comprising an rib gene cluster encoding the E. coli O25B antigen, wherein the rib gene cluster encodes:
- a. dTDP-Glucose 4,6-dehydratase;
- b. dTDP-6-Deoxy-D-glucose 3,5-epimerase;
- c. Glucose-1-phosphate thymidylyltransferase;
- d. dTDP-4-dehydrorhamnose 3,5-epimerase;
- e. O antigen flippase;
- f. dTDP-Rha:Glc-Rha(Ac)-GlcNAc-UPP α-1,3-rhamnosyltransferase;
- g. UDP-Glc:Glc-Rha(Ac)-GlcNAc-UPP β-1,6-glucosyltransferase;
- h. O antigen polymerase;
- i. O-acetyl transferase;
- j. UDP-Glc:Rha-GlcNAc-UPP α-1,3-glucosyltransferase;
- and
- k. dTDP-Rha:
- GlcNAc-UPP α-1,3-rhamnosyltransferase;
- ii. a nucleotide sequence encoding an oligosaccharyl transferase;
- and
- iii. a nucleotide sequence encoding the carrier protein, wherein the carrier protein comprises a consensus sequence Asn-X-Ser(Thr), and X is any natural amino acid except Pro (SEQ ID NO:
- 14);
- and
- (b) purifying the bioconjugate.
Dependencies are inferred from the claim text.