Oligosaccharide modification and labeling of proteins
The present invention generally relates to methods of functionalizing proteins, particularly antibodies, at oligosaccharide linkages, methods of humanizing antibodies by modifying glycosylation, as well as to novel antibodies linked to modified oligosaccharides. The invention further relates to kits that may be used to produce the antibodies of the invention.
1. An antibody comprising a labeled oligosaccharide, wherein the labeled oligosaccharide comprises:
an azido-modified sugar attached to a GlcNAc residue on the antibody and the azido-modified sugar is N-azidogalactosamine (GalNAz); and
a reporter molecule, a carrier molecule or a solid support comprising an activated alkyne group capable of reacting with said azido-modified sugar;
wherein said reporter molecule, carrier molecule or solid support attaches to the azido group of the azido-modified sugar through the activated alkyne group.
2. The antibody of claim 1 , wherein the antibody is humanized.
3. The antibody of claim 1 , wherein the antibody is a therapeutic antibody.
4. The antibody of claim 3 , wherein the reporter molecule is a fluorescent dye, enzyme, radiolabel, a metal chelator, or a detectable substrate.
5. The antibody of claim 1 , wherein the label is selected from the group consisting of therapeutic moieties, therapeutic agents, radioactive metal ions, DNA, protein, peptides, sugars, detectable labels, biotin, and avidin.
6. The antibody of claim 1 , wherein the antibody is obtained using a method comprising:
attaching an azido-modified sugar to a GlcNAc residue on the antibody using a mutant β-1,4-galactosyl transferase (GalT) enzyme, wherein the azido-modified sugar is N-azidogalactosamine (GalNAz); and
mixing said antibody with a reporter molecule, carrier molecule or solid support comprising an activated alkyne group capable of reacting with said azido group of the GalNAz;
wherein said reporter molecule, carrier molecule or solid support attaches to the antibody at said azido group of the GalNAz through said activated alkyne group, thereby forming the antibody comprising the labeled oligosaccharide.