3-galactosyl-N-acetylglucosaminide 4-alpha-L-fucosyltransferase FUT3
4-galactosyl-N-acetylglucosaminide 3-alpha-L-fucosyltransferase activity
Distinct human α(1,3)-fucosyltransferases drive Lewis-X/sialyl Lewis-X assembly in human cells.
In humans, six α(1,3)-fucosyltransferases (α(1,3)-FTs: FT3/FT4/FT5/FT6/FT7/FT9) reportedly fucosylate terminal lactosaminyl glycans yielding Lewis-X (Le<sup>X</sup>; CD15) and/or sialyl Lewis-X (sLe<sup>X</sup>; CD15s), structures that play key functions in cell migration, development, and immunity. Prior studies analyzing α(1,3)-FT specificities utilized either purified and/or recombinant enzymes to modify synthetic substrates under nonphysiological reaction conditions or molecular biology approaches wherein α(1,3)-FTs were expressed in mammalian cell lines, notably excluding investigations using primary human cells. Accordingly, although significant insights into α(1,3)-FT catalytic properties have been obtained, uncertainty persists regarding their human Le<sup>X</sup>/sLe<sup>X</sup> biosynthetic range across various glycoconjugates. Here, we undertook a comprehensive evaluation of the lactosaminyl product specificities of intracellularly expressed α(1,3)-FTs using a clinically relevant primary human cell type, mesenchymal stem cells. Cells were transfected with modified mRNA encoding each human α(1,3)-FT, and the resultant α(1,3)-fucosylated lactosaminyl glycoconjugates were analyzed using a combination of flow cytometry and MS. The data show that biosynthesis of sLe<sup>X</sup> is driven by FTs-3, -5, -6, and -7, with FT6 and FT7 having highest potency. FT4 and FT9 dominantly biosynthesize Le<sup>X</sup>, and, among all FTs, FT6 holds a unique capacity in creating sLe<sup>X</sup> and Le<sup>X</sup> determinants across protein and lipid glycoconjugates. Surprisingly, FT4 does not generate sLe<sup>X</sup> on glycolipids, and neither FT4, FT6, nor FT9 synthesizes the internally fucosylated sialyllactosamine VIM-2 (CD65s). These results unveil the relevant human lactosaminyl glycans created by human α(1,3)-FTs, providing novel insights on how these isoenzymes stereoselectively shape biosynthesis of vital glycoconjugates, thereby biochemically programming human cell migration and tuning human immunologic and developmental processes.