N-Linked Glycoengineering Service in Plant Cell

N-Glycosylation in Plant and Mammalian Cells

There are differences in N-glycosylation between plant and mammalian systems, especially in the structure of complex glycans. Plant cells typically synthesize a limited repertoire of N-glycan structures, including the prevalent GnGnXF structures, characterized by β1,2-linked xylose and α1,3-linked fucose. Paucimannosidic MMXF structures synthesized by β-N-acetylhexosaminidases (HEXO) are also common in plants. In certain cases, plant proteins may exhibit Lea structures, characterized by terminal β1,3-galactosylation and α1,4-fucosylation. In contrast, mammalian N-glycans do not contain xylose residues, and α1,6-fucose is attached to the proximal GlcNAc. Mammalian N-glycan characterized by the attachment of β1,4 galactose is absent in plant N-glycans. These distinctions emphasize the necessity for N-linked glycoengineering in plant-based expression systems. Remarkably, the limited glycosylation capacity of plants has proven advantageous for producing proteins with homogeneous glycans.

Fig.1 N-glycosylation in plant and mammalian cells.^{1, 3}

N-Linked Glycoengineering Services in Plant Cell at Creative Biolabs

By employing cellular engineering techniques such as knockout/knockdown and overexpression of glycosylation enzymes, Plant Cell Glycoengineering Services have been successfully developed and are readily accessible at Creative Biolabs. For N-linked glycoengineering in plant cells, we have devised two main types of modifications. The first is the elimination of plant-specific N-glycans to ensure the homogeneity of recombinant glycoproteins. The second is the introduction of human-like glycosylation pathways, which enables the production of glycoproteins with glycans that mimic the native human glycosylation patterns, resulting in a more biologically relevant product. By combining these strategies, plant expression systems can be customized to yield glycoproteins with desired human-type glycan structures, suitable for various applications.

• Elimination of unwanted plant-specific N-glycans
• Elimination of XylT and FUT11/12 to inhibit the attachment of β1,2-xylose and α1,3-fucose.
• Elimination of HEXOs to increase complex N-glycans with terminal GlcNAc residues.
• Elimination of GalT1 and FUT1 to inhibit the synthesis of the Lea epitopes.
• Introduction of human N-glycans
• Introduction of core α1,6-fucose by expression of human α1,6-fucosyltransferase.
• Introduction of N-glycans containing β1,4 galactose by expressing human β1,4-GalT.
• Introduction of bisecting GlcNAc by expressing human GnT-III.
• Introduction of multi-antennary complex glycans by expressing human GnT-IV/V.
• Introduction of genes responsible for mammalian sialylation pathway.

Advantages of Our Services

• Highly efficient and functional knockdown and knockout
• High-level expression vector for overexpression
• Multiple and comprehensive strategies of glycoengineering
• Extensive experience in cell line glycoengineering

Published Data

Technology: Plant cell glycoengineering

Journal: Frontiers in Plant Science

IF: 6.627

Published: 2014

Results: The researcher utilized several methods involving the knock-out or knock-down of plant-specific β1,2-XT and core α1,3-FT and introduction of GnTIV and GnTV, β1,4-GalT, and the biosynthetic pathway for sialylation, to transform the glycans present in Nicotiana benthamiana wild-type plants into a glycosylation profile that resembles human serum Erythropoietin.

Fig.2 N-glycoengineering in plants to produce tetra-sialylated proteins.^{2, 3}

Creative Biolabs offers comprehensive plant glycoengineering services, allowing for tailored manipulation of glycosylation processes. For further details or specific requirements, please don't hesitate to contact us.

References

1. Schoberer, Jennifer, and Richard Strasser. "Plant glyco-biotechnology." Seminars in Cell & Developmental Biology. Vol. 80. Academic Press, 2018.
2. Loos, Andreas, and Herta Steinkellner. "Plant glyco-biotechnology on the way to synthetic biology." Frontiers in plant science 5 (2014): 523.

3. Under Open Access license CC BY 4.0, without modification.

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• O-Linked Glycoengineering in Plant Cell