Molecular cloning and functional expression of beta1, 2-xylosyltransferase cDNA from Arabidopsis thaliana.
The transfer of xylose from UDP-xylose to the core beta-linked mannose of N-linked oligosaccharides by beta1,2-xylosyltransferase (XylT) is a widespread feature of plant glycoproteins which renders them immunogenic and allergenic in man. Here, we report the isolation of the Arabidopsis thaliana XylT gene, which contains two introns and encodes a 60.2 kDa protein with a predicted type II transmembrane protein topology typical for Golgi glycosyltransferases. Upon expression of A. thaliana XylT cDNA in the baculovirus/insect cell system, a recombinant protein was produced that exhibited XylT activity in vitro. Furthermore, the recombinant enzyme displayed XylT activity in vivo in the insect cells, as judged by the acquired cross-reaction of cellular glycoproteins with antibodies against the beta1,2-xylose epitope. The cloned XylT cDNA as well as the recombinant enzyme are essential tools to study the role of beta1,2-xylose in the immunogenicity and allergenicity of plant glycoproteins at the molecular level.
Strasser
R
R
Zentrum für Angewandte Genetik, Universität für Bodenkultur Wien, Muthgasse 18, A-1190, Vienna, Austria.
Mucha
J
J
Mach
L
L
Altmann
F
F
Wilson
I B
IB
Glössl
J
J
Steinkellner
H
H
D000595:Amino Acid Sequence; D000818:Animals; D017360:Arabidopsis / Q000235:genetics*; D001483:Base Sequence; D002460:Cell Line; D003001:Cloning, Molecular; D018076:DNA, Complementary; D008969:Molecular Sequence Data; D010430:Pentosyltransferases / Q000235:genetics* / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D018411:Spodoptera / Q000166:cytology
D018076:DNA, Complementary; D011994:Recombinant Proteins; D010430:Pentosyltransferases; C408586:beta 1,2-xylosyltransferase