Production of human milk oligosaccharides in microbial hosts with engineered import / export
AssigneeJennewein Biotechnologie GmbH
InventorsStefan JENNEWEIN, Dirk WARTENBERG
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Abstract
The present invention relates to methods for the production of oligosaccharides in genetically modified bacterial host cells, as well as to the genetically modified host cells used in the methods. The genetically modified host cell comprises at least one recombinant glycosyltransferase, and at least one nucleic acid sequence coding for a protein enabling the export of the oligosaccharide.
Claims23
A method for production of an oligosaccharide by a genetically modified microbial host cell, comprising
- providing a genetically modified microbial host cell that comprises:
- at least one recombinant glycosyltransferase;
- increased expression or activity of at least one homologous or heterologous sugar export protein capable of exporting the oligosaccharide;
- and
- decreased expression or inactivation of at least one export protein that exports precursors of the oligosaccharide from the host cell;
- cultivating the host cell in a medium under conditions permissive for the production of the oligosaccharide, whereby the oligosaccharide is exported into the medium at an increased level compared to the unmodified host cell, and
- obtaining the oligosaccharide from the medium.
A genetically modified microbial host cell for the production of a oligosaccharide, the oligosaccharide comprising a lacto-N-triose II (LNT-II;
- GlcNAc(β1-3)Gal(β1-4)Gluc) as a core trisaccharide, wherein the host cell comprises
- at least one recombinant glycosyltransferase, the glycosyltransferase being optionally selected from a galactosyltransferase, a sialyltransferase, and an N-acetylglucosaminyltransferase;
- at least one homologous or heterologous nucleic acid sequence coding for a sugar export protein capable of exporting of the oligosaccharide into a culture medium the host cell is cultivated in, wherein said sugar export protein capable of exporting the oligosaccharide belongs to the class of secondary active transporters, and wherein said homologous or heterologous nucleic acid sequence coding for the sugar export protein capable of exporting the oligosaccharide is overexpressed or under control of a promoter sequence enabling the overexpression of said nucleic acid sequence;
- and
- inactivated expression or activity of at least one endogenous sugar export protein for export of a precursor of the oligosaccharide.
Dependencies are inferred from the claim text.