Identification and analysis of cell wall glycan epitopes and polyphenol oxidase in pawpaw (Asimina triloba [L.] Dunal) fruit pulp as affected by high pressure processing and refrigerated storage.

This research explores the cell wall composition and polyphenol oxidase activity of two pawpaw (Asimina triloba) fruit varieties, Susquehanna and Green River Belle, that were subjected to high pressure processing and 45 days of refrigerated storage. We hypothesize that high pressure processing may inhibit enzymatic action responsible for pawpaw's deleterious postharvest tissue softening and browning. Glycome profiling uses mAb groupings that recognize 19 groups of glycan epitopes present in most major classes of cell wall glycans and was used to determine cell wall composition. Results show that both varieties have typical type I primary cell walls of flowering dicots. However, differences in the fine cell wall structure between the varieties can be inferred and the varieties behaved differently during refrigerated storage, likely indicating of a difference in cell wall-modifying enzymes present in the primary cell walls. High pressure processing treatment does not seem to be effective at eliminating polyphenol oxidase activity.

Arabidopsis thaliana RGXT1 and RGXT2 encode Golgi-localized (1,3)-alpha-D-xylosyltransferases involved in the synthesis of pectic rhamnogalacturonan-II.

Two homologous plant-specific Arabidopsis thaliana genes, RGXT1 and RGXT2, belong to a new family of glycosyltransferases (CAZy GT-family-77) and encode cell wall (1,3)-alpha-d-xylosyltransferases. The deduced amino acid sequences contain single transmembrane domains near the N terminus, indicative of a type II membrane protein structure. Soluble secreted forms of the corresponding proteins expressed in insect cells showed xylosyltransferase activity, transferring d-xylose from UDP-alpha-d-xylose to l-fucose. The disaccharide product was hydrolyzed by alpha-xylosidase, whereas no reaction was catalyzed by beta-xylosidase. Furthermore, the regio- and stereochemistry of the methyl xylosyl-fucoside was determined by nuclear magnetic resonance to be an alpha-(1,3) linkage, demonstrating the isolated glycosyltransferases to be (1,3)-alpha-d-xylosyltransferases. This particular linkage is only known in rhamnogalacturonan-II, a complex polysaccharide essential to vascular plants, and is conserved across higher plant families. Rhamnogalacturonan-II isolated from both RGXT1 and RGXT2 T-DNA insertional mutants functioned as specific acceptor molecules in the xylosyltransferase assay. Expression of RGXT1- and RGXT2-enhanced green fluorescent protein constructs in Arabidopsis revealed that both fusion proteins were targeted to a Brefeldin A-sensitive compartment and also colocalized with the Golgi marker dye BODIPY TR ceramide, consistent with targeting to the Golgi apparatus. Taken together, these results suggest that RGXT1 and RGXT2 encode Golgi-localized (1,3)-alpha-d-xylosyltransferases involved in the biosynthesis of pectic rhamnogalacturonan-II.