Disclosed are genetically engineered microbial cells for the production of oligosaccharides comprising a galactose-β1,4-glucose moiety at their reducing end, wherein said microbial cells are able to produce said oligosaccharides in the absence of exogenously added lactose, and a method of producing said oligosaccharides using said microbial cells.

The present invention relates to a process for the enzymatic modification of a glycoprotein. The process comprises the step of contacting a glycoprotein comprising a glycan comprising a terminal GlcNAc-moiety, in the presence of glycosyltransferase that is, or is derived from, a β-(1,4)-N-acetylgalactosaminyltransferase, with a non-natural sugar-derivative nucleotide. The non-natural sugar-derivative nucleotide is according to formula (3):

##STR00001##

wherein A is selected from the group consisting of —N.sub.3, —C(O)R.sup.3, —(CH.sub.2).sub.iC≡C—R.sup.4, —SH, —SC(O)R.sup.8, —SC(O)OR.sup.8, —SC(S)OR.sup.8, —F, —Cl, —Br —I, —OS(O).sub.2R.sup.5, terminal C.sub.2-C.sub.24 alkenyl groups, C.sub.3-C.sub.5 cycloalkenyl groups, C.sub.4-C.sub.8 alkadienyl groups, terminal C.sub.3-C.sub.24 allenyl groups and amino groups. The invention further relates to a glycoprotein obtainable by the process according to the invention, to a bioconjugate that can be obtained by conjugating the glycoprotein with a linker-conjugate, and to β-(1,4)-N-acetylgalactosaminyltransferases that can be used in preparing the glycoprotein according to the invention.

Methods for preparing hypersialylated IgG are described.

This disclosure relates to glycoengineering, and methods of utilizing glycoengineering for various therapeutic purposes.

The present invention relates to a process for the modification of a glycoprotein, using a β-(1,4)-N-acetylgalactosaminyltransferase or a mutant thereof. The process comprises the step of contacting a glycoprotein comprising a glycan comprising a terminal GlcNAc-moiety, in the presence of a β-(1,4)-N-acetylgalactosaminyl-transferase or a mutant thereof, with anon-natural sugar-derivative nucleotide. The non-natural sugar-derivative nucleotideis according to formula (3), wherein A is selected from the group consisting of —N.sub.3; —C(0)R.sup.3; —C═C—R.sup.4; —SH; —SC(0)R.sup.8; —SC(V)OR.sup.8, wherein V is O or S; —X wherein X is selected from the group consisting of F, Cl, Br and I; —OS(0).sub.2R.sup.5; an optionally substituted C.sub.2-C.sub.24 alkyl group; an optionally substituted terminal C.sub.2-C.sub.24 alkenyl group; and an optionally substituted terminal C.sub.3-C.sub.24 alkenyl group.

Described herein are fusion proteins, e.g., fusion proteins comprising enzymatically active portion(s) of ST6Gall or B4GalT1 as well as methods for producing them, nucleic acid molecule(s) encoding the fusion protein(s), vectors comprising the nucleic acid molecule(s), and host cell(s) comprising the vector(s). Also described herein are methods of sialyating immunoglobulin G (IgG) antibodies.

Herein is reported a method for the enzymatic production of an antibody with a modified glycosylation in the Fc-region comprising the steps of incubating the antibody light chain affinity ligand-bound monoclonal antibody with a glycosylation in the Fc-region with a first enzyme for a time sufficient and under conditions suitable to modify the glycosylation of the Fc-region, recovering the antibody from the antibody light chain affinity ligand, incubating the recovered antibody in solution with a second enzyme for a time sufficient and under conditions suitable to modify the glycosylation of the Fc-region to a defined form, separating the antibody with the modified glycosylation in the Fc-region from the second enzyme in a cation exchange chromatography, and thereby producing the antibody with a modified glycosylation in the Fc-region.

The disclosure discloses genetically engineered bacteria producing lacto-N-neotetraose and a production method thereof, and belongs to the fields of metabolic engineering and food biotechnology. To solve the problem of low yield of lacto-N-neotetraose produced by a microbial method in the prior art, through exogenous expression of lgtA and lgtB, reasonable combination and regulation of overexpression of lacY, pgm, galE, galT and galK in a lacto-N-neotetraose synthesis pathway, knockout of lacZ expression in an Escherichia coli host, and optimization of a carbon source in the culture process, the disclosure achieves the objectives of regulating the carbon flux of a metabolic pathway and improving the yield of lacto-N-neotetraose. In a shake flask experiment, the yield of lacto-N-neotetraose produced by E. coli increased from 304 mg/L to 1031 mg/L, laying a foundation for industrial production of the lacto-N-neotetraose.

Disclosed herein are methods galatosylating IgG antibodies, methods of preparing hypersialylated (hsIgG), e.g., using immobilized β1,4-Galactosyltransferase I (β4GalT1), as well as polypeptides comprising β1,4-Galactosyltransferase I (β4GalT1) bound to a solid support and compositions comprising the same.

Disclosed herein are dual drug antibody drug conjugates with defined stoichiometric ratios of each drug. The conjugates disclosed herein are useful for the treatment of cancer, particularly drug resistant and multidrug resistance cancer.