Described herein are compositions and methods of producing glycosylated proteins in vitro and in vivo. The methods include using host cells to produce glycosylated proteins. Also described herein are glycosylated proteins produced using such methods and uses thereof.

Methods of modulating the properties of a cell culture expressing a protein of interest are provided. In various embodiments the methods relate to the overexpression of proteins involved in the N-glycosylation pathway.

Methods of modulating the properties of a cell culture expressing a protein of interest are provided. In various embodiments the methods relate to the overexpression of proteins involved in the N-glycosylation pathway.

The present disclosure relates to a nucleic acid construct having a chimeric nucleic acid molecule encoding a tripartite glycosyltransferase fusion protein. The chimeric nucleic acid molecule includes a first nucleic acid moiety encoding an amphipathic shield domain protein; a second nucleic acid moiety encoding a glycosyltransferase; and a third nucleic acid moiety encoding a water soluble expression decoy protein. The first nucleic acid moiety is coupled to the second nucleic acid moiety's 3 end and the third nucleic acid moiety is coupled to the second nucleic acid moiety's 5 end. The coupling may be direct or indirect. The present disclosure further relates to an expression vector, a host cell, and a tripartite glycosyltransferase fusion protein encoded by the nucleic acid construct. Also disclosed are methods of recombinantly producing a tripartite glycosyltransferase fusion protein in soluble form and methods of cell-free glycan remodeling.

A quadruple mutant (qm) plant is deficient of functions of core 1,3-fucosyltransferase A (FucTA), core 1,3-fucosyltransferase B (FucTB), 1,2-xylosyltransferase (XylT), and 1,2-N-acetylglucosaminyltransferase II (GnTII), and produces a protein containing hypoallergenic pauci-mannose-type N-glycan that does not include 1,3-fucose and 1,2-xylose residues. A method for producing a transgenic plant for a production of a protein containing hypoallergenic pauci-mannose-type N-glycan that does not include 1,3-fucose and 1,2-xylose residues but includes 1,6-fucose residue includes preparing the quadruple mutant (qm) plant, and transforming the quadruple mutant plant with a recombinant vector containing a gene encoding the human-derived 1,6-fucosyltransferase (FUT8) protein to overexpress FUT8 gene, and selecting a transgenic plant which is deficient of the functions of FucTA, FucTB, XylT and GnTII proteins.

The present disclosure relates to recombinant proteins having N-acetylglucosaminyltransferase activity. The present disclosure further relates to methods for producing complex N-glycans including the steps of providing host cells containing such recombinant proteins and culturing the host cells such that the recombinant proteins are expressed.

Provided herein are host cells capable of producing a human milk oligosaccharide (HMO), such as yeast cells that include one or more heterologous nucleic acids encoding one or more enzymes of the HMO biosynthetic pathway, such as a fucosyltransferase, GDP-mannose dehydratase, lactose permease, and/or fucose synthase. Also provided are fermentation compositions including the disclosed host cells, as well as related methods of producing and recovering HMOs generated by the host cells.

The present invention relates to a cell-free enzyme-catalyzed process for producing glycoproteins of general formula (I) from a lipid-linked oligosaccharide and a peptide. Further, said process includes the construction of the lipid-linked oligosaccharide from a mannose trisaccharide containing core structure. Particularly, the lipid-linked oligosaccharide is a high mannose-, complex-, or hybrid-type N-glycan.