IP Library › Granted Patent US 12,351,806
Granted Patent B2
US 12,351,806 · App. 17/759,261 · Granted Jul 8, 2025

HMO production

Inventors: Manos Papadakis (Brønshøj, DK); Katrine Bych Kampmann (Valby, DK)
Assignee: GLYCOM A/S
C12N15/70C12P19/02
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Quick Facts
Patent No.
US 12,351,806
App. No.
17/759,261
Granted
Jul 8, 2025
Kind
B2
Abstract

The present inventive concept relates to a genetically modified cell enabled for the production of an oligosaccharide, preferably, an HMO, comprising a recombinant nucleic acid encoding a protein of the MFS superfamily; and methods using said cell for the production the oligosaccharide, preferably an HMO.

Claims (25)

1. A genetically modified cell capable of producing one or more Human Milk Oligosaccharides (HMOs) under fermentative conditions, wherein the cell comprises a recombinant nucleic acid encoding a protein of SEQ ID NO: 1, or a functional homologue thereof which amino acid sequence is at least 90% identical to SEQ ID NO: 1,

wherein the functional homologue increases the total production of the one or more HMOs while reducing by-product formation or facilitates the export of the one or more HMOs out of the genetically modified cell.

2. The genetically modified cell according to claim 1 , wherein the one or more HMOs is selected from the group consisting of 2′-fucosyllactose (2′FL), 3′-fucosyllactose (3FL), difucosyllactose (DFL), 3′-sialyllactose (3′SL), 6′-sialyllactose (6′SL), Lacto-N-Triose-2 (LNT-2), Lacto-N-neotetraose (LNnT), Lacto-N-tetraose (LNT), Lacto-N-fucopentaose I (LNFP-I), Lacto-N-fucopentaose II (LNFP-II), Lacto-N-fucopentaose III (LNFP-III), Lacto-N-fucopentaose IV (LNFP-IV), and Lacto-N-fucopentaose V (LNFP-V), and/or para-lacto-N-neohexaose (pLNnH); or a mixture thereof.

3. The genetically modified cell according to claim 1 , wherein the genetically modified cell is Escherichia coli.

4. The genetically modified cell according to claim 1 , wherein the cell further comprises an expression element comprising a lac promoter or a glp promoter.

5. The genetically modified cell according to claim 4 , wherein the lac promoter, if present, is Plac and the glp promoter, if present, is Pg/pF.

6. A nucleic acid construct comprising a nucleic acid sequence encoding a protein of SEQ ID NO: 1, or a functional homologue thereof, having more than 90% sequence identity to SEQ ID NO: 1, wherein the nucleic acid sequence encoding a protein of SEQ ID NO: 1, has at least 80% sequence identity to SEQ ID NO: 2.

7. The nucleic acid construct according to claim 6 , wherein the construct further comprises a nucleic acid sequence comprising an expression element comprising a lac promoter or a glp promoter.

8. The nucleic acid construct according to claim 7 , wherein the expression element regulates the expression of the nucleic acid sequence having at least 85% sequence identity to SEQ ID NO: 2.

9. The nucleic acid construct according to claim 7 , wherein the lac promoter, if present, is Plac and the glp promoter, if present, is Pg/pF.

10. A method for the production of one or more HMOs, the method comprising the steps of:

(i) providing the genetically modified cell of claim 1 ;

(ii) culturing the cell according to (i) in a suitable cell culture medium to express said recombinant nucleic acid, whereby one or more HMOs are produced by the cultured genetically modified cell;

(iii) harvesting the one or more HMOs produced in step (ii).

11. The method according to claim 10 , wherein the one or more HMOs is selected from the group consisting of 2′-FL, 3-FL, DLF, LNT, LNT-II, LNnT, pLNH-II and pLNnH; or a mixture thereof.

12. The method according to claim 10 , wherein the one or more HMOs is selected from the group consisting of LNT, LNT-II, LNnT, and pLNH-II and pLNnH; or a mixture thereof.

13. The genetically modified cell according to claim 1 , wherein the functional homologue comprises an amino acid sequence at least 95% identical to SEQ ID NO: 1.

14. The method according to claim 10 , wherein the one or more HMOs is selected from the group consisting of 2′-fucosyllactose (2′FL), 3′-fucosyllactose (3FL), difucosyllactose (DFL), 3′-sialyllactose (3′SL), 6′-sialyllactose (6′SL), Lacto-N-Triose-2 (LNT-2), Lacto-N-neotetraose (LNnT), Lacto-N-tetraose (LNT), Lacto-N-fucopentaose I (LNFP-I), Lacto-N-fucopentaose II (LNFP-II), Lacto-N-fucopentaose III (LNFP-III), Lacto-N-fucopentaose IV (LNFP-IV), and Lacto-N-fucopentaose V (LNFP-V), and/or para-lacto-N-neohexaose (pLNnH); or a mixture thereof.

15. A method for the production of one or more HMOs, the method comprising the steps of:

(i) providing the genetically modified cell of claim 4 ;

(ii) culturing the cell according to (i) in a suitable cell culture medium to express said recombinant nucleic acid, whereby one or more HMOs are produced by the cultured genetically modified cell;

(iv) harvesting one or more HMOs produced in step (ii).

16. The method according to claim 15 , wherein the one or more HMOs is selected from the group consisting of 2′-fucosyllactose (2′FL), 3′-fucosyllactose (3FL), difucosyllactose (DFL), 3′-sialyllactose (3′SL), 6′-sialyllactose (6′SL), Lacto-N-Triose-2 (LNT-2), Lacto-N-neotetraose (LNnT), Lacto-N-tetraose (LNT), Lacto-N-fucopentaose I (LNFP-I), Lacto-N-fucopentaose II (LNFP-II), Lacto-N-fucopentaose III (LNFP-III), Lacto-N-fucopentaose IV (LNFP-IV), and Lacto-N-fucopentaose V (LNFP-V), and/or para-lacto-N-neohexaose (pLNnH); or a mixture thereof.

17. The method according to claim 15 , wherein the one or more HMOs is selected from the group consisting of 2′-FL, 3-FL, DLF, LNT, LNT-II, LNnT, pLNH-II and pLNnH; or a mixture thereof.

18. The method according to claim 15 , wherein the one or more HMOs is selected from the group consisting of LNT, LNT-II, LNnT, and pLNH-II and pLNnH; or a mixture thereof.

Assignments (1)
ASSIGNMENT OF ASSIGNOR'S INTEREST Recorded Jul 19, 2023
From: PAPADAKIS, MANOS; KAMPMANN, KATRINE BYCH
To: GLYCOM A/S
Reel/Frame 064314/0639 →
Priority Claims (1)
DK PA 2020 00087 · Jan 23, 2020 · national
Continuity (1)
Related Publication 20230046359A1 · Feb 16, 2023
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