Acceptor specificity of the human leukocyte alpha3 fucosyltransferase: role of FucT-VII in the generation of selectin ligands

The alpha3 fucosyltransferase, FucT-VII, is one of the keyglycosyltransferases involved in the biosynthesis of the sialyl Lewis X(sLex) antigen on human leukocytes. The sialyl Lewis X antigen(NeuAcalpha(2-3)Galbeta(1-4)[Fucalpha(1-3)]GlcNAc-R) is an essentialcomponent of the recruitment of leukocytes to sites of inflammation,mediating the primary interaction between circulating leukocytes andactivated endothelium. In order to characterize the enzymatic properties ofthe leukocyte alpha3 fucosyltransferase FucT-VII, the enzyme has beenexpressed in Trichoplusia ni insect cells. The enzyme is capable ofsynthesizing both sLexand sialyl-dimeric-Lexstructures in vitro , from3'-sialyl-lacNAc and VIM-2 structures, respectively, with only low levelsof fucose transfer observed to neutral or 3'-sulfated acceptors. Studiesusing fucosylated NeuAcalpha(2-3)-(Galbeta(1- 4)GlcNAc)3-Me acceptorsdemonstrate that FucT-VII is able to synthesize both di-fucosylated andtri-fucosylated structures from mono- fucosylated precursors, butpreferentially fucosylates the distal GlcNAc within a polylactosaminechain. Furthermore, the rate of fucosylation of the internal GlcNAcresidues is reduced once fucose has been added to the distal GlcNAc. Theseresults indicate that FucT-VII is capable of generating complex selectinligands, in vitro , however the order of fucose addition to the lactosaminechain affects the rate of selectin ligand synthesis.