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US 2007/0178551 A1Published application
Methods for producing modified glycoproteins
AssigneeGlycoFi, Inc.
InventorTillman Gerngross
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AssigneeGlycoFi, Inc.
InventorTillman Gerngross
Estimating the term…
Cell lines having genetically modified glycosylation pathways that allow them to carry out a sequence of enzymatic reactions, which mimic the processing of glycoproteins in humans, have been developed. Recombinant proteins expressed in these engineered hosts yield glycoproteins more similar, if not substantially identical, to their human counterparts. The lower eukaryotes, which ordinarily produce high-mannose containing N-glycans, including unicellular and multicellular fungi are modified to produce N-glycans such as Man5GlcNAc2 or other structures along human glycosylation pathways. This is achieved using a combination of engineering and/or selection of strains which: do not express certain enzymes which create the undesirable complex structures characteristic of the fungal glycoproteins, which express exogenous enzymes selected either to have optimal activity under the conditions present in the fungi where activity is desired, or which are targeted to an organelle where optimal activity is achieved, and combinations thereof wherein the genetically engineered eukaryote expresses multiple exogenous enzymes required to produce “human-like” glycoproteins.
1-8. (canceled)
A method for producing a recombinant glycoprotein comprising an N-glycan structure that comprises a GlcNAcMan5GlcNAc2 glycoform in a lower eukaryotic host cell selected or genetically modified to produce N-glycan structures having an excess of 30 mole % of a Man5GlcNAc2 glycoform that can serve as a substrate for GlcNAc transferase I in vivo, the method comprising the step of introducing into said host cell a nucleic acid encoding a GlcNAc transferase I enzyme selected to have optimal activity in the ER or Golgi of said host cell, the enzyme comprising:
A method for producing a recombinant glycoprotein comprising an N-glycan comprising a GlcNAcMan3GlcNAc2 glycoform in a lower eukaryotic host cell selected or genetically modified to produce N-glycan structures having an excess of 30 mole % of a Man5GlcNAc2 glycoform that are converted in vivo to a GlcNAcMan5GlcNAc2 glycoform by GlcNAc transferase I activity localized in the ER or Golgi apparatus of the host cell, the method comprising the step of introducing into the host cell a nucleic acid encoding a mannosidase II enzyme selected to have optimal activity in the ER or Golgi of said host cell, the enzyme comprising:
A method for producing a recombinant glycoprotein comprising an N-glycan comprising a GlcNAc2Man3GlcNAc2 glycoform in a lower eukaryotic host cell selected or genetically modified to produce N-glycan structures having an excess of 30 mole % of a Man5GlcNAc2 glycoform that are converted in vivo to a GlcNAcMan3GlcNAc2 glycoform by GlcNAc transferase I and mannosidase II localized in the ER or Golgi apparatus of the host cell, the method comprising the step of introducing into the host cell a nucleic acid encoding a GlcNAc transferase II enzyme selected to have optimal activity in the ER or Golgi of said host cell, the enzyme comprising:
Dependencies are inferred from the claim text.