Fucosylation-deficient cells
AssigneeRegeneron Pharmaceuticals, Inc.
InventorsGang Chen, Darya Burakov, James P. Fandl
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Abstract
An isolated nucleic acid encoding an FX protein having a serine at position 79, a lysine at position 90, a leucine at position 136, an arginine at position 211, a serine at position 289, and a combination thereof is provided. Cells having a gene encoding a modified FX protein are provided, wherein the cells exhibit a reduced ability to fucosylate a glycoprotein at a first temperature, but exhibit the ability to fucosylate the glycoprotein at a second temperature. Methods and compositions for making glycoproteins with reduced fucosylation are provided.
Claims13
A method for making a glycoprotein with reduced fucosylation, comprising:
- culturing a mammalian cell that expresses a GDP-4-keto-6-deoxymannose-3,5-epimerase-4-reductase (FX) protein comprising the amino acid sequence of SEQ ID NO:
- 1, except for an amino acid substitution of L289S, at a temperature of about 37° C. and in the absence of an external fucose source, wherein the cell expresses a glycoprotein;
- and, isolating the glycoprotein from the culture.
A method for making a glycoprotein with reduced fucosylation, comprising:
- culturing a mammalian cell that expresses a FX protein that comprises the amino acid sequence of SEQ ID NO:
- 1, except for a first amino acid substitution of L289S and a second amino acid substitution selected from the group consisting of N79S, N90K, P136L, and G211R, at a temperature of about 37° C. and in the absence of an external fucose source, wherein the cell expresses a glycoprotein;
- and, isolating the glycoprotein from the culture.
Dependencies are inferred from the claim text.