Construction of a functional CMP-sialic acid biosynthesis pathway in Arabidopsis.
Previous studies have reported that plants contain negligible amounts of free or protein-bound N-acetylneuraminic acid (Neu5Ac). This is a major disadvantage for the use of plants as a biopharmaceutical expression system, since N-glycans with terminal Neu5Ac residues are important for the biological activities and half-lives of recombinant therapeutic glycoproteins in humans. For the synthesis of Neu5Ac-containing N-glycans, plants have to acquire the ability to synthesize Neu5Ac and its nucleotide-activated derivative, cytidine monophospho-N-acetylneuraminic acid. In this study, we have generated transgenic Arabidopsis (Arabidopsis thaliana) plants expressing three key enzymes of the mammalian Neu5Ac biosynthesis pathway: UDP-N-acetylglucosamine 2-epimerase/N-acetylmannosamine kinase, N-acetylneuraminic acid phosphate synthase, and CMP-N-acetylneuraminic acid synthetase. Simultaneous expression of UDP-N-acetylglucosamine 2-epimerase/N-acetylmannosamine kinase and N-acetylneuraminic acid phosphate synthase resulted in the generation of significant Neu5Ac amounts (1,275 nmol g(-1) fresh weight in leaves) in planta, which could be further converted to cytidine monophospho-N-acetylneuraminic acid (2.4 nmol g(-1) fresh weight in leaves) by coexpression of CMP-N-acetylneuraminic acid synthetase. These findings are a major step toward the production of Neu5Ac-containing glycoproteins in plants.
Castilho
Alexandra
A
Institute of Applied Genetics and Cell Biology , University of Natural Resources and Applied Life Sciences, BOKU-Vienna, A-1190 Vienna, Austria.
Pabst
Martin
M
Leonard
Renaud
R
Veit
Christiane
C
Altmann
Friedrich
F
Mach
Lukas
L
Glössl
Josef
J
Strasser
Richard
R
Steinkellner
Herta
H
D000818:Animals; D017360:Arabidopsis / Q000201:enzymology* / Q000235:genetics; D002238:Carbohydrate Epimerases / Q000378:metabolism; D003569:Cytidine Monophosphate N-Acetylneuraminic Acid / Q000378:metabolism; D006801:Humans; D051379:Mice; D019158:N-Acetylneuraminic Acid / Q000096:biosynthesis*; D000216:N-Acylneuraminate Cytidylyltransferase / Q000378:metabolism*; D007652:Oxo-Acid-Lyases / Q000378:metabolism*; D030821:Plants, Genetically Modified / Q000201:enzymology; D011134:Polysaccharides / Q000378:metabolism
D011134:Polysaccharides; D003569:Cytidine Monophosphate N-Acetylneuraminic Acid; C018767:N-acylneuraminate-9-phosphate synthase; D000216:N-Acylneuraminate Cytidylyltransferase; D007652:Oxo-Acid-Lyases; D002238:Carbohydrate Epimerases; C100033:UDP acetylglucosamine-2-epimerase; D019158:N-Acetylneuraminic Acid