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Microorganism for producing human milk oligosaccharide

A technology of human milk oligosaccharides and microorganisms, which is applied in the field of purifying HMO and preparing HMO, can solve the problems of difficulty in obtaining purity and the cost of non-robust HMO.

Pending Publication Date: 2020-05-01
OLIGOSCI BIOTECH GMBH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, in both cases it was difficult to obtain large quantities and / or achieve satisfactory purity
[0005] HMOs can also be chemically synthesized, however, this technique is not robust and the cost of chemically synthesized HMOs is high

Method used

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  • Microorganism for producing human milk oligosaccharide
  • Microorganism for producing human milk oligosaccharide
  • Microorganism for producing human milk oligosaccharide

Examples

Experimental program
Comparison scheme
Effect test

Embodiment approach

[0108] Genetically modified microorganisms for the preparation of HMOs

[0109] The present invention provides genetically modified microorganisms for use in the production of HMOs.

[0110] The HMO prepared in the present invention is based on the disaccharide lactose or N-acetyllactosamine (N-acetyllactosamine). Preferably, the HMO is lactose based.

[0111] Preferred human milk oligosaccharides in the context of the present invention include, but are not limited to, 2'-fucosyllactose (2-FL), 3'-fucosyllactose (3-FL), 3'-sialic acid Lactose (3-SL), 6'-sialyllactose (6-SL), lacto-N-tetraose (LNT), lacto-N-neotetraose (LNnT), lacto-N-hexaose (LNH), Iso-lacto-N-octaose, iso-lacto-N-neo-octaose, p-lacto-N-octaose, lacto-N-fucopentose I (LNFP I), lacto-N-fucopentose II (LNFP II), lacto-N-fucopentose III (LNFP III), lacto-N-fucopentose V (LNFP V), LS-tetrasaccharide a (LST a), LS-hexasaccharide b ( LST b), LS-hexaose c (LST c) and disialyllacto-N-tetraose (DSLNT).

[0112] In...

Embodiment

[0175] Materials and methods

[0176] The strains, plasmids and primers used are listed in Tables 3 and 4. Escherichia coli BL21(DE3) was used as the host strain for HMO production, and Escherichia coli TOP10 was used for plasmid construction and maintenance. Escherichia coli in the presence of ampicillin (100 μg ml -1 ), streptomycin (50 μg ml -1 ) or kanamycin (50 μg ml -1 ) of LB medium (10gL -1 Tryptone, 5gL -1 Yeast extract, 10gL -1 cultured in NaCl). For gene deletions, the λRed recombinase / flipse system from Jensen et al. 2015 can be used (Sci Rep 2015; 5:17874). The vector pSIJ8 was transformed into competent cells of Escherichia coli BL21(DE3). The lambda Red gene was induced in the presence of 15 mM L-arabinose. The kanamycin resistance expression cassette (kanR) was amplified from vector pKD13 (Datsenko and Wanner, 2000) flanked by a 50 bp long homologous target sequence and FRT sites, and transformed into L-arabinose-induced receptive in state cells. Fol...

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Abstract

Human milk oligosaccharides (HMOs) may be used e.g. as functional ingredients in infant nutrition, medical nutrition, functional foods and animal feed. There is still a need of improved means of producing HMOs. The present invention provides genetically modified microorganisms for the improved production of HMOs and HMO production methods using the same. The microorganisms of the invention may have one or more yield-enhancing modifications, including an inducible lysis system, which allows for the easy extraction of intracellular and extracellular HMOs, lactose permease mutants, which may increase intracellular lactose levels, or chaperones, which may increase intracellular availability of key enzymes for the production of HMOs.

Description

technical field [0001] The present invention relates to genetically modified microorganisms for producing human milk oligosaccharides (HMOs) and methods for producing HMOs using said microorganisms. The present invention also relates to a medium for cultivating the microorganism of the present invention, a purified HMO prepared by the preparation method of the present invention, and uses of the medium and the HMO. Background technique [0002] Human milk oligosaccharides (HMOs) are structurally diverse unconjugated glycans found in human milk. More than 200 HMOs have been identified so far, all of which are composed of only five monosaccharide structural units, namely D-galactose (Gal), D-glucose (Glc), N-acetyl-D-glucosamine (GlcNAc ), L-fucose (Fuc) and sialic acid derivative N-acetyl-neuraminic acid, and lactose moiety coupled to construct (Petschacher and Nidetzky, J Biotechnol.2016,235:61-83). The reducing end can be lactose or lactose extended by several disaccharide...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K14/245C07K14/435C12P19/18C12N9/10A23L33/00
CPCC07K14/245C07K14/435C12P19/18C12N9/1051A23L33/00Y02A50/30A23K20/163C12N1/20C12Y204/0104
Inventor M·佩拉查S·维姆霍夫F·迈因哈特
Owner OLIGOSCI BIOTECH GMBH