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Abstract
A. thaliana has thirteen FUTs, ten of which are classified as members of glycosyltransferase family 37 (GT37) according to the Carbohydrate-Active enZYmes (CAZy) database, and all of them are predicted to be Golgi-localized type-II transmembrane proteins (Lombard et al., 2014; Sarria et al., 2001). Thus far, three out of the ten GT37 FUTs from A. thaliana have been functionally characterized: AtFUT1, AtFUT4, and AtFUT6. AtFUT1 has been extensively studied biologically, biochemically, and structurally, and has been demonstrated to be xyloglucan-specific, and has been established as the sole FUT carrying out this function in planta. AtFUT4 and AtFUT6 have been less extensively studied but have been previously suggested to be partially or non-redundant, arabinogalactan protein (AGP) specific FUTs. Further and more detailed study of fusion-protein versions of AtFUT4 and AtFUT6 produced by transient transfection in mammalian HEK293 cell lines, has confirmed the AGP specificity of AtFUT4 and AtFUT6 and further lead to novel discoveries on their activities and specificities. Numerous and varied in vitro assays, have demonstrated that AtFUT4 and AtFUT6 are in fact identical in their recognition and ability to fucosylate various arabinogalactan (AG) and non-AG-like oligosaccharides, proving that they are in fact enzymatically redundant. Additionally, their ability to recognize and fucosylate various non-AG oligosaccharides, further confirms the fucosylation site as residing on an arabinose residue but extends the breadth of oligo- and polysaccharides that these enzymes can recognize and successfully fucosylate. In planta studies on the ATFUT4 and ATFUT6 genes utilizing Promoter: β-glucuronidase expression and localization has demonstrated that in the A. thaliana root, the only tissue in which both ATFUT4 and ATFUT6 co-localize, ATFUT4 sub-localizes to the elongation zone, while ATFUT6 sub-localizes to the meristematic zone. These differences in sub-localization, serve as a possible explanation as to why both ATFUT4 and ATFUT6 are functional in the root, despite carrying out the same exact function enzymatically. Additionally, here we report the first study on the activities of some of the remaining members of the GT37 family, demonstrating that some exhibit FUT-specific activity towards the hydrolysis of GDP-Fuc.