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38 results about "Endoglycosidase" patented technology
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An Endoglycosidase is an enzyme that releases oligosaccharides from glycoproteins or glycolipids. It may also cleave polysaccharide chains between residues that are not the terminal residue, although releasing oligosaccharides from conjugated protein and lipid molecules is more common.
The present invention is aimed to provide a method for preparing an acceptor that is N-glycan hydrolyzed antibody or a Fc fragment thereof used for producing antibody having a homogeneous N-glycan structure; a method for determining a combination of endoglycosidases for use in said preparation; and a method for measuring N-glycans linked to an antibody. The present invention is directed to a method for producing a N-glycan hydrolyzed antibody or Fc fragment thereof, comprising reacting the antibody or the Fc fragment thereof with several endoglycosidases; and a method for determining quantitative information of an objective N-glycan with a desired structure linked to an antibody or a Fc thereof, comprising a protease treatment step and a glycopeptide measurement step, etc.
The invention belongs to the technical field of analytical chemistry, and particularly relates to a method for catalyzing an isotope labeled N-sugar chain by using endoglycosidase. Glycoprotein enzymolysis reaction of the endoglycosidase is performed in 18O water, and when the N-sugar chain is digested by enzyme, an isotope 18O molecule is labeled at the reductive tail end of the N-sugar chain. Mass spectrometric detection results prove that the difference between the isotope 18O labeled sugar chain molecular weight and the unlabeled sugar chain molecular weight is 2 Daltons and the isotope 18O molecule is stably labeled at the reductive tail end of the sugar chain. The method is simple and efficient, fills the blank of the sugar chain isotope 18O labeling technology, and can be effectively used for relative quantification of the sugar chain.
The invention discloses a method for detecting an activity of an endoglycosidase. The method includes providing the endoglycosidase with a substrate of the endoglycosidase and detecting cleavage of the substrate. The method further includes at least partly inhibiting the transglycosidase activity of the endoglycosidase. The transglycosidase activity may be inhibited by chemically modifying the substrate such that transglycosylation of the substrate by the endoglycosidase is at least partly inhibited while the endoglycosidase is still capable of cleaving the substrate. In one embodiment, the substrate comprises an oligosaccharide chain. Compounds and kits suitable for use in a method of the invention are also disclosed. Methods involving competitive inhibitors are also disclosed as are methods for the synthesis of glycosylated substrates involving the transglycosidase activity of an endoglycosidase.
The present invention is aimed to provide a method for preparing an acceptor that is N-glycan hydrolyzed antibody or a Fc fragment thereof used for producing antibody having a homogeneous N-glycan structure; a method for determining a combination of endoglycosidases for use in said preparation; and a method for measuring N-glycans linked to an antibody. The present invention is directed to a method for producing a N-glycan hydrolyzed antibody or Fc fragment thereof, comprising reacting the antibody or the Fc fragment thereof with several endoglycosidases; and a method for determining quantitative information of an objective N-glycan with a desired structure linked to an antibody or a Fc thereof, comprising a protease treatment step and a glycopeptide measurement step, etc.
Genetically engineered host animal cells capable of producing glycoproteins having modified glycosylation patterns, e.g., defucosylation and / or monoglycosylation. Such host animal cells can be engineered to express fucosidase, endoglycosidase or both.
A mutant of Endos2 includes one or more mutations in the sequence of a wild-type EndoS2 (SEQ ID NO:1), wherein the one or more mutations are in a peptide region located within residues 133-143, residues 177-182, residues 184-189, residues 221-231, and / or residues 227-237, wherein the mutant of EndoS2 has a low hydrolyzing activity and a high tranglycosylation activity, as compared to those of the wild-type EndoS2. A method for preparing an engineered glycoprotein using the mutant of EndoS2 includes coupling an activated oligosaccharide to a glycoproteinacceptor. The activated oligosaccharide is a glycanoxazoline.
The invention discloses a method for detecting an activity of an endoglycosidase. The method includes providing the endoglycosidase with a substrate of the endoglycosidase and detecting cleavage of the substrate. The method further includes at least partly inhibiting the transglycosidase activity of the endoglycosidase. The transglycosidase activity may be inhibited by chemically modifying the substrate such that transglycosylation of the substrate by the endoglycosidase is at least partly inhibited while the endoglycosidase is still capable of cleaving the substrate. In one embodiment, the substrate comprises an oligosaccharide chain. Compounds and kits suitable for use in a method of the invention are also disclosed. Methods involving competitive inhibitors are also disclosed as are methods for the synthesis of glycosylated substrates involving the transglycosidase activity of an endoglycosidase.
Genetically engineered host animal cells are capable of producing glycoproteins with modified glycosylation patterns, such as defucosylation and / or monoglycosylation. Such host animal cells can be engineered to express fucosidases, endoglycosidases, or both.
A mutant of EndoS2 includes one or more mutations in the sequence of a wild-type EndoS2 (SEQ ID NO: 1), wherein the one or more mutations are in a peptide region located within residues 133-143, residues 177-182, residues 184-189, residues 221-231, and / or residues 227-237, wherein the mutant of EndoS2 has a low hydrolyzing activity and a high tranglycosylation activity, as compared to those of thewild-type EndoS2. A method for preparing an engineered glycoprotein using the mutant of EndoS2 includes coupling an activated oligosaccharide to a glycoproteinacceptor. The activated oligosaccharideis a glycanoxazoline.