Arabidopsis thaliana beta1,2-xylosyltransferase: an unusual glycosyltransferase with the potential to act at multiple stages of the plant N-glycosylation pathway.
XylT (beta1,2-xylosyltransferase) is a unique Golgi-bound glycosyltransferase that is involved in the biosynthesis of glycoprotein-bound N-glycans in plants. To delineate the catalytic domain of XylT, a series of N-terminal deletion mutants was heterologously expressed in insect cells. Whereas the first 54 residues could be deleted without affecting the catalytic activity of the enzyme, removal of an additional five amino acids led to the formation of an inactive protein. Characterization of the N-glycosylation status of recombinant XylT revealed that all three potential N-glycosylation sites of the protein are occupied by N-linked oligosaccharides. However, an unglycosylated version of the enzyme displayed substantial catalytic activity, demonstrating that N-glycosylation is not essential for proper folding of XylT. In contrast with most other glycosyltransferases, XylT is enzymatically active in the absence of added metal ions. This feature is not due to any metal ion directly associated with the enzyme. The precise acceptor substrate specificity of XylT was assessed with several physiologically relevant compounds and the xylosylated reaction products were subsequently tested as substrates of other Golgi-resident glycosyltransferases. These experiments revealed that the substrate specificity of XylT permits the enzyme to act at multiple stages of the plant N-glycosylation pathway.
Bencúr
Peter
P
Department für Angewandte Pflanzenwissenschaften und Pflanzenbiotechnologie, Institut für Angewandte Genetik und Zellbiologie, Universität für Bodenkultur Wien, Muthgasse 18, A-1190 Wien, Austria.
Steinkellner
Herta
H
Svoboda
Barbara
B
Mucha
Jan
J
Strasser
Richard
R
Kolarich
Daniel
D
Hann
Stephan
S
Köllensperger
Gunda
G
Glössl
Josef
J
Altmann
Friedrich
F
Mach
Lukas
L
D000818:Animals; D017360:Arabidopsis / Q000201:enzymology*; D020134:Catalytic Domain; D002460:Cell Line; D003067:Coenzymes; D015870:Gene Expression; D006031:Glycosylation; D006056:Golgi Apparatus / Q000201:enzymology; D008670:Metals; D010430:Pentosyltransferases / Q000378:metabolism*; D013379:Substrate Specificity
D003067:Coenzymes; D008670:Metals; D010430:Pentosyltransferases; C408586:beta 1,2-xylosyltransferase