Distinct substrate specificities of five human alpha-1,3-fucosyltransferases for in vivo synthesis of the sialyl Lewis x and Lewis x epitopes.
Five different human alpha-1,3-fucosyltransferase genes, the Fuc-TIII, Fuc-TIV, Fuc-TV, Fuc-TVI and Fuc-TVII genes, have been cloned to date. We transfected HeLa cells and Namalwa cells with each of the five different genes, and established a series of stable cloned transformant cells. Thin-layer chromatography immunostaining analysis revealed that all five enzymes were able to synthesize sialyl Lewis x (sLe(x)) epitopes on glycolipids in HeLa cells, but each enzyme showed a different preference as to the carbohydrate chain length on glycolipids as acceptor substrates. Fuc-TIII and Fuc-TV showed very similar patterns of sLe(x) positive bands, which indicated that the enzymes have similar acceptor substrate specificities. Fuc-TVI exhibited a little different pattern from those of the former two enzymes. Fuc-TIV and Fuc-TVII showed similarity in the positive bands, however, their patterns were quite different from those of the former three enzymes. Four enzymes except for Fuc-TVII were able to synthesize the Lewis x (Le(x)) epitope on glycolipids in HeLa cells. Fuc-TV alone showed a little different pattern of Le(x) positive bands from those of the other three enzymes. Flow cytometric analysis of HeLa cells and Namalwa cells again demonstrated the similar specificities of Fuc-TIII and Fuc-TV. They exhibited similar stronger staining with FH6 (anti-sLe(x)) antibodies than that with the other enzymes. A phylogenetic tree of the five enzymes constructed using the neighbor-joining method showed good agreement with the similarities in the enzyme substrate specificity.
Kimura
H
H
Division of Cell Biology, Institute of Life Science, Soka University, Tokyo, Japan.
Shinya
N
N
Nishihara
S
S
Kaneko
M
M
Irimura
T
T
Narimatsu
H
H
D002051:Burkitt Lymphoma; D002999:Clone Cells; D005434:Flow Cytometry; D005647:Fucosyltransferases / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D007527:Isoenzymes / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D016256:Lewis X Antigen / Q000096:biosynthesis*; D009844:Oligosaccharides / Q000096:biosynthesis*; D011994:Recombinant Proteins / Q000378:metabolism; D000080506:Sialyl Lewis X Antigen; D013379:Substrate Specificity; D014158:Transcription, Genetic; D014162:Transfection
D007527:Isoenzymes; D016256:Lewis X Antigen; D009844:Oligosaccharides; D011994:Recombinant Proteins; D000080506:Sialyl Lewis X Antigen; D005647:Fucosyltransferases; C031692:galactoside 3-fucosyltransferase