Provided herein are genetically modified yeast cells capable of producing one or more human milk oligosaccharides. The yeast cells include one or more heterologous nucleic acids that encode enzymes of a human milk oligosaccharide biosynthetic pathway. The yeast cells do not include a heterologous nucleic acid encoding a fucokinase. Also provided are fermentation compositions including the disclosed genetically modified yeast cells, and related methods of producing and recovering human milk oligosaccharides generated by the yeast cells.

The present invention provides a novel biosynthetic production process which converts L-galactose into 2-fucosyllactose via four enzymatically catalyzed reaction steps. The present process is designed such that co-factors required by the process are regenerated within the four reaction steps, hence making the process cost-effective and efficient. The process can be performed in vitro in a cell-free system. The present invention also provides mutant enzymes that can be used to increase production levels of 2-fucosyllactose, whether using the novel pathway described herein or the mannose-dependent pathway known in the art.

The present invention relates to a method for increasing the productivity of 2-fucosyllactose through various changes in culture medium composition and culturing on the basis of lactose, which is a substrate, wherein 2-fucosyllactose can be continuously produced in a high-yield at an optimum lactose concentration discovered by a culturing method of the present invention.

Microbial cells genetically engineered with a heterologous nucleic acid sequence that increases export of 2 fucosyllactose are disclosed. Methods of increasing export of 2 fucosyllactose from a microbial cell and for identifying a heterologous nucleic acid sequence that increases export of 2 fucosyllactose from a microbial cell are also disclosed.

The present invention relates to a genetically modified cell expressing an -1,2-fucosyltransferase and an -1,3-fucosyltransferase, and a transporter protein of the major facilitator superfamily (MFS) and to a method for recombinant production of human milk oligosaccharides (HMOs) using said genetically modified cell. More particularly, the invention provides a method for recombinant production of and a genetically modified cell capable of producing difucosyllactose (DFL) as the most abundant HMO, with a relatively low content of 3-fucosyllactose (3FL) and/or 2-fucosyllactose (2FL).

The invention provides modular cell-free de-novo synthesis of glycans with immobilized bionanocatalysts. The invention provides materials, and in particular, magnetic materials, for producing glycans of defined length and sequences using one or more enzymes that are immobilized within bionanocatalysts (BNCs) which in turn are embedded within scaffolds to control the synthesis in batch or continuous processes manufacturing. In some embodiments, the scaffolds are high magnetism and high porosity composite blends of thermoplastics or thermosets comprising magnetic particles that form powders. In some embodiments, Selective Laser Sintering (SLS) is used to design and produce objects via 3D printing by sintering composite magnetic powders. The modular flow cells may be mixed and matched for a highly customizable and highly efficient cell-free manufacturing process. In some embodiments the elementary and system modules provided by the invention are employed. In preferred embodiments, human milk oligosaccharides (HMOs) are produced.

Microbial cells genetically engineered with a heterologous nucleic acid sequence that increases export of 2 fucosyllactose are disclosed. Methods of increasing export of 2 fucosyllactose from a microbial cell and for identifying a heterologous nucleic acid sequence that increases export of 2 fucosyllactose from a microbial cell are also disclosed.

The present invention is in the technical field of synthetic biology and metabolic engineering. More particularly, the present invention is in the technical field of metabolically engineered cells and use of said cell in a cultivation, preferably a fermentation. The present invention describes a cell for the production of a compound. The cell comprises a pathway for the production of the compound, which can be a disaccharide, oligosaccharide and/or a Neu(n)Ac-containing bioproduct, wherein (n) is 4, 5, 7, 8 or 9 or a combination thereof. The cell is metabolically engineered for enhanced synthesis of acetyl-Coenzyme A. The invention also resides in a method of producing such compound by cultivation, preferably a fermentation, with such a cell.

The invention provides compositions and methods for engineering E. coli or other host production bacterial strains to produce fucosylated oligosaccharides, and the use thereof in the prevention or treatment of infection.

An enzyme that it is a fusion protein. The enzyme includes an N-terminal domain of at least amino acids 1-129 of SEQ ID No. 7 or an amino acid sequence at least 80% identical thereto and that includes at least amino acids 155-286 of SEQ ID No. 5 or an amino acid sequence at least 80% identical thereto as a C-terminal domain and has fucosyltransferase activity. The N-terminal domain and the C-terminal domain being derived from two different fucosyltransferases.