title: Arabinoxylan biosynthesis in wheat. Characterization of arabinosyltransferase activity in Golgi membranes.
abstract: Arabinoxylan arabinosyltransferase (AX-AraT) activity was investigated using microsomes and Golgi vesicles isolated from wheat (Triticum aestivum) seedlings. Incubation of microsomes with UDP-[(14)C]-beta-L-arabinopyranose resulted in incorporation of radioactivity into two different products, although most of the radioactivity was present in xylose (Xyl), indicating a high degree of UDP-arabinose (Ara) epimerization. In isolated Golgi vesicles, the epimerization was negligible, and incubation with UDP-[(14)C]Ara resulted in formation of a product that could be solubilized with proteinase K. In contrast, when Golgi vesicles were incubated with UDP-[(14)C]Ara in the presence of unlabeled UDP-Xyl, the product obtained could be solubilized with xylanase, whereas proteinase K had no effect. Thus, the AX-AraT is dependent on the synthesis of unsubstituted xylan acting as acceptor. Further analysis of the radiolabeled product formed in the presence of unlabeled UDP-Xyl revealed that it had an apparent molecular mass of approximately 500 kD. Furthermore, the total incorporation of [(14)C]Ara was dependent on the time of incubation and the amount of Golgi protein used. AX-AraT activity had a pH optimum at 6, and required the presence of divalent cations, Mn(2+) being the most efficient. In the absence of UDP-Xyl, a single arabinosylated protein with an apparent molecular mass of 40 kD was radiolabeled. The [(14)C]Ara labeling became reversible by adding unlabeled UDP-Xyl to the reaction medium. The possible role of this protein in arabinoxylan biosynthesis is discussed.
authors.affiliation: Plant Biochemistry Laboratory, Department of Plant Biology, The Royal Veterinary and Agricultural University, 1871 Frederiksberg C, Copenhagen, Denmark.
authors.forename: Andrea Celia
authors.initials: AC
authors.lastname: Porchia
authors.forename: Susanne Oxenbøll
authors.initials: SO
authors.lastname: Sørensen
authors.forename: Henrik Vibe
authors.initials: HV
authors.lastname: Scheller
mesh_terms: D002250:Carbon Radioisotopes; D002850:Chromatography, Gel; D006056:Golgi Apparatus / Q000201:enzymology / Q000378:metabolism*; D007425:Intracellular Membranes / Q000187:drug effects / Q000378:metabolism; D010430:Pentosyltransferases / Q000378:metabolism*; D014908:Triticum / Q000187:drug effects / Q000201:enzymology* / Q000378:metabolism; D014539:Uridine Diphosphate Sugars / Q000494:pharmacology; D014990:Xylans / Q000096:biosynthesis*; D014994:Xylose / Q000378:metabolism
chemical_list: D002250:Carbon Radioisotopes; D014539:Uridine Diphosphate Sugars; D014990:Xylans; C040817:uridine diphosphate arabinose; C085118:arabinoxylan; D014994:Xylose; D010430:Pentosyltransferases; C101967:arabinosyltransferase