title: Efficient metabolic oligosaccharide engineering of glycoproteins by UDP-N-acetylglucosamine 2-epimerase/N-acetylmannosamine kinase (GNE) knock-down.
abstract: Improving the accessibility and functions of therapeutic and diagnostic glycoproteins is one of the major goals of glycobiotechnology. Here we present that stable knock-down of UDP-N-acetylglucosamine 2-epimerase/N-acetylmannosamine kinase (GNE), the key enzyme in the sialic acid biosynthetic pathway, dramatically increases incorporation of N-acetylmannosamine analogues into glycoproteins of HEK293 cells. By means of these GNE-deficient cells highly sialylated glycoproteins can efficiently be decorated with reactive functional groups, which can be employed in bioorthogonal functionalization strategies for fluorescence labelling or biotinylation.
authors.affiliation: Beuth Hochschule für Technik Berlin (University of Applied Sciences), Department of Life Sciences and Technology, Seestrasse 64, 13347 Berlin, Germany.
authors.forename: Heinz
authors.initials: H
authors.lastname: Möller
authors.forename: Verena
authors.initials: V
authors.lastname: Böhrsch
authors.forename: Lothar
authors.initials: L
authors.lastname: Lucka
authors.forename: Christian P R
authors.initials: CP
authors.lastname: Hackenberger
authors.forename: Stephan
authors.initials: S
authors.lastname: Hinderlich
mesh_terms: D015703:Antigens, CD / Q000378:metabolism; D002236:Carbohydrate Conformation; D002238:Carbohydrate Epimerases / Q000378:metabolism; D015815:Cell Adhesion Molecules / Q000378:metabolism; D055785:Gene Knockdown Techniques; D006023:Glycoproteins / Q000737:chemistry / Q000378:metabolism*; D057809:HEK293 Cells; D006801:Humans; D019158:N-Acetylneuraminic Acid / Q000378:metabolism; D009844:Oligosaccharides / Q000737:chemistry / Q000378:metabolism*; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000172:deficiency / Q000378:metabolism*; D015202:Protein Engineering / Q000379:methods*; D034741:RNA, Small Interfering / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism; D013194:Staining and Labeling
chemical_list: D015703:Antigens, CD; C069847:CD66 antigens; D015815:Cell Adhesion Molecules; D006023:Glycoproteins; D009844:Oligosaccharides; D034741:RNA, Small Interfering; D011994:Recombinant Proteins; D017853:Phosphotransferases (Alcohol Group Acceptor); C018765:N-acylmannosamine kinase; D002238:Carbohydrate Epimerases; C100033:UDP acetylglucosamine-2-epimerase; D019158:N-Acetylneuraminic Acid