Sialyiation of N-linked glycoproteins in the baculovirus expression vector system
AssigneeBoyce Thompson Institute for Plant Research, Inc.
InventorH. Alan Wood
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Abstract
The present disclosure utilizes a novel approach to protein preparation in the baculovirus expression vector system (BEVS). Specifically, the present invention analyzes the effects of microgravity/low shear on complex glycosylation of proteins prepared via BEVS, including the addition of terminal sialic acid residues to N-linked oligosaccharides.
Claims12
A method of expressing a recombinant gene in an insect cell line by using a baculovirus expression system such that said recombinant gene being expressed in said baculovirus expression system codes for a protein with N-linked glycosylation, comprising the steps of;
- a) culturing cells for use in said baculovirus expression system;
- b) infecting said cells with a recombinant baculovirus that expresses said recombinant gene encoding a recombinant protein that would normally have N-linked glycosylation if expressed in the source organism of said recombinant gene;
- and
- c) culturing said infected cells in a microgravity environment, in a medium that includes serum.
A method of expressing a recombinant gene in an insect cell line by using a baculovirus expression system such that said recombinant gene being expressed in said baculovirus expression system codes for a protein with N-linked glycosylation, comprising the steps of;
- a) culturing cells for use in said baculovirus expression system;
- b) infecting said cells with a recombinant baculovirus that expresses said recombinant gene encoding a recombinant protein that would normally have oligosaccharides with terminal sialic acid residues during N-linked glycosylation if expressed in the source organism of said recombinant gene;
- and
- c) culturing said infected cells in a microgravity environment, in a medium that includes serum, such that said protein with N-linked glycosylation obtains oligosaccharides having terminal sialic acid residues.
A method of culturing insect cells comprising the steps of:
- a) culturing insect cells in a medium that includes serum;
- and
- b) culturing said cells in a microgravity environment, such that N-acetylglucosaminyl-transferase and galactosyl-transferase activities are induced.
A method of culturing insect cells comprising the steps of:
- culturing cells from a cell line designated BTI-Tn-5B1-4h in a microgravity environment and a medium that includes serum, such that N-acetylglucosaminyl-transferase, galactosyl-transferase or sialic acid transferase activity is induced.
A cell line designated BTI-Tn-5B1-4h, characterized in that N-acetylglucosaminyl-transferase, galactosyl-transferase and sialic acid transferase activities are induced when said cell line is cultured in medium that includes serum.
Dependencies are inferred from the claim text.