Production of Multi-Antennary N-Glycan Structures in Plants
The invention provides methods for producing multi-antennary glycoproteins in plant and plant cells. In particular the invention provides plants comprising a chimeric gene comprising glucosaminyltransferase IV and plants comprising two chimeric genes comprising glucosaminyltransferase IV and V.
1 . A method to produce multi-antennary glycoproteins in plant cells comprising the steps of:
a. providing a plant cell with a chimeric gene comprising the following operably linked nucleic acid molecules:
i) a plant-expressible promoter,
ii) a DNA region encoding a functional N-acetylglucosaminyltransferase IV,
iii) a DNA region involved in transcription termination and polyadenylation, and
b. cultivating said plant cell and isolating multi-antennary glycoproteins from said plant cell.
2 . The method according to claim 1 further comprising the step of providing the plant cell with a second chimeric gene comprising the following operably linked nucleic acid molecules:
i) a plant expressible promoter,
ii) a DNA region encoding a functional N-acetylglucosaminyltransferase V,
iii) a DNA region involved in transcription termination and polyadenylation.
3 . The method according to claim 1 wherein the plant cells have no detectable beta-(1,2)xylosyltransferase and no detectable alfa-(1,3)fucosyltransferase activity.
4 . The method according to claim 1 wherein said N-acetylglucosaminyltransferase IV and/or said N-acetylglucosaminyltransferase V are of the mammalian type.
5 . The method according to claim 1 wherein said N-acetylglucosaminyltransferase IV and/or said N-acetylglucosaminyltransferase V are of the hybrid type.
6 . The method according to claim 1 further introducing a third chimeric gene comprising a plant expressible promoter and a DNA region encoding a beta(1,4)-galactosyltransferase.
7 . The method according to claim 1 wherein a heterologous glycoprotein is additionally expressed in said plant cells from a chimeric gene comprising a plant expressible promoter and a DNA region encoding said heterologous glycoprotein.
8 . A multi-antennary glycoprotein obtained by the method of claim 1 .
9 . A plant cell comprising a chimeric gene comprising the following operably linked nucleic acid molecules:
i) a plant-expressible promoter,
ii) a DNA region encoding a functional N-acetylglucosaminyltransferase IV,
iii) a DNA region involved in transcription termination and polyadenylation.
10 . The plant cell according to claim 9 further comprising a second chimeric gene comprising the following operably linked nucleic acid molecules:
i) a plant expressible promoter,
ii) a DNA region encoding a functional N-acetylglucosaminyltransferase V,
iii) a DNA region involved in transcription termination and polyadenylation.
11 . The plant cell according to claim 9 wherein said plant cell has no detectable beta-(1,2)xylosyltransferase activity and no detectable alfa-(1,3)fucosyltransferase activity.
12 . The plant cell according to claim 9 wherein said N-acetylglucosaminyltransferase IV and/or said N-acetylglucosaminyltransferase V are of the mammalian type.
13 . The plant cell according to claim 9 wherein said N-acetylglucosaminyltransferase IV and/or said N-acetylglucosaminyltransferase V are of the hybrid type.
14 . The plant cell according to claim 9 comprising a heterologous glycoprotein which is expressed in said plant cell from a chimeric gene comprising a plant expressible promoter and a DNA region encoding said heterologous glycoprotein.
15 . A plant consisting essentially of the plant cell according to claim 9 .