Molecular cloning and characterization of a novel alpha 1,2-fucosyltransferase (CE2FT-1) from Caenorhabditis elegans.
Here we report the discovery of a unique fucosyltransferase (FT) in Caenorhabditis elegans. In studying the activities of FTs in extracts of adult C. elegans, we detected activity toward the unusual disaccharide acceptors Galbeta1-4Xyl-R and Galbeta1-6GlcNAc-R to generate products with the general structure Fucalpha1-2Galbeta1-R. We identified a gene encoding a unique alpha1,2FT (designated CE2FT-1), which contains an open reading frame encoding a predicted protein of 355 amino acids with the type 2 topology and domain structure typical of other glycosyltransferases. The predicted cDNA for CE2FT-1 has very low identity (5-10%) at the amino acid level to alpha1,2FT sequences in humans, rabbits, and mice. Recombinant CE2FT-1 expressed in human 293T cells has high alpha1,2FT activity toward the simple acceptor Galbeta-O-phenyl acceptor to generate Fucalpha1-2Galbeta-R, which in this respect resembles mammalian alpha1,2FTs. However, CE2FT-1 is otherwise completely different from known alpha1,2FTs in its acceptor specificity, since it is unable to fucosylate either Galbeta1-4Glcbeta-R or free lactose and prefers the unusual acceptors Galbeta1-4Xylbeta-R and Galbeta1-6GlcNAc-R. Promoter analysis of the CE2FT-1 gene using green fluorescent protein reporter constructs demonstrates that CE2FT-1 is expressed in single cells of early stage embryos and exclusively in the 20 intestinal cells of L(1)-L(4) and adult worms. These and other results suggest that multiple fucosyltransferase genes in C. elegans may encode enzymes with unique activities, expression, and developmental roles.
Zheng
Qinlong
Q
Department of Biochemistry and Molecular Biology, University of Oklahoma Health Sciences Center, Oklahoma City, Oklahoma 73104, USA.
Van Die
Irma
I
Cummings
Richard D
RD
D000595:Amino Acid Sequence; D000818:Animals; D030801:Animals, Genetically Modified; D001483:Base Sequence; D015153:Blotting, Western; D017173:Caenorhabditis elegans / Q000201:enzymology* / Q000235:genetics*; D003001:Cloning, Molecular; D004247:DNA / Q000378:metabolism; D018076:DNA, Complementary / Q000378:metabolism; D005647:Fucosyltransferases / Q000737:chemistry* / Q000235:genetics*; D017930:Genes, Reporter; D049452:Green Fluorescent Proteins; D006801:Humans; D008164:Luminescent Proteins / Q000378:metabolism; D051379:Mice; D008954:Models, Biological; D008969:Molecular Sequence Data; D010957:Plasmids / Q000378:metabolism; D011401:Promoter Regions, Genetic; D017434:Protein Structure, Tertiary; D012333:RNA, Messenger / Q000378:metabolism; D011817:Rabbits; D011994:Recombinant Proteins / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017386:Sequence Homology, Amino Acid; D013552:Swine; D000097631:Galactoside 2-alpha-L-fucosyltransferase
D004247:DNA; D018076:DNA, Complementary; D005647:Fucosyltransferases; D049452:Green Fluorescent Proteins; D008164:Luminescent Proteins; D012333:RNA, Messenger; D011994:Recombinant Proteins; D000097631:Galactoside 2-alpha-L-fucosyltransferase