Dataset Information

Molecular cloning of pigeon UDP-galactose:beta-D-galactoside alpha1,4-galactosyltransferase and UDP-galactose:beta-D-galactoside beta1,4-galactosyltransferase, two novel enzymes catalyzing the formation of Gal alpha1-4Gal beta1-4Gal beta1-4GlcNAc sequence.

ABSTRACT: We previously found that pigeon IgG possesses unique N-glycan structures that contain the Gal alpha1-4Gal beta1-4Gal beta1-4GlcNAc sequence at their nonreducing termini. This sequence is most likely produced by putative alpha1,4- and beta1,4-galactosyltransferases (GalTs), which are responsible for the biosynthesis of the Gal alpha1-4Gal and Gal beta1-4Gal sequences on the N-glycans, respectively. Because no such glycan structures have been found in mammalian glycoproteins, the biosynthetic enzymes that produce these glycans are likely to have distinct substrate specificities from the known mammalian GalTs. To study these enzymes, we cloned the pigeon liver cDNAs encoding alpha4GalT and beta4GalT by expression cloning and characterized these enzymes using the recombinant proteins. The deduced amino acid sequence of pigeon alpha4GalT has 58.2% identity to human alpha4GalT and 68.0 and 66.6% identity to putative alpha4GalTs from chicken and zebra finch, respectively. Unlike human and putative chicken alpha4GalTs, which possess globotriosylceramide synthase activity, pigeon alpha4GalT preferred to catalyze formation of the Gal alpha1-4Gal sequence on glycoproteins. In contrast, the sequence of pigeon beta4GalT revealed a type II transmembrane protein consisting of 438 amino acid residues, with no significant homology to the glycosyltransferases so far identified from mammals and chicken. However, hypothetical proteins from zebra finch (78.8% identity), frogs (58.9-60.4%), zebrafish (37.1-43.0%), and spotted green pufferfish (43.3%) were similar to pigeon beta4GalT, suggesting that the pigeon beta4GalT gene was inherited from the common ancestors of these vertebrates. The sequence analysis revealed that pigeon beta4GalT and its homologs form a new family of glycosyltransferases.

SUBMITTER: Suzuki N

PROVIDER: S-EPMC2820745 | biostudies-literature | 2010 Feb

REPOSITORIES: biostudies-literature

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Molecular cloning of pigeon UDP-galactose:beta-D-galactoside alpha1,4-galactosyltransferase and UDP-galactose:beta-D-galactoside beta1,4-galactosyltransferase, two novel enzymes catalyzing the formation of Gal alpha1-4Gal beta1-4Gal beta1-4GlcNAc sequence.

Suzuki Noriko N Yamamoto Kazuo K

The Journal of biological chemistry 20091203 8

We previously found that pigeon IgG possesses unique N-glycan structures that contain the Gal alpha1-4Gal beta1-4Gal beta1-4GlcNAc sequence at their nonreducing termini. This sequence is most likely produced by putative alpha1,4- and beta1,4-galactosyltransferases (GalTs), which are responsible for the biosynthesis of the Gal alpha1-4Gal and Gal beta1-4Gal sequences on the N-glycans, respectively. Because no such glycan structures have been found in mammalian glycoproteins, the biosynthetic enzy ...[more]

PMID: 19959475

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Project description:The localization and activity of the enzyme UDP-galactose-hydroxy fatty acid-containing ceramide galactosyltransferase is described in rat brain myelin subfractions during development. Other lipid-synthesizing enzymes, such as cerebroside sulphotransferase, UDP-glucose-ceramide glucosyltransferase and CDP-choline-1,2-diacylglycerol cholinephosphotransferase, were also studied for comparison in myelin subfractions and microsomal membranes. The purified myelin was subfractionated by isopycnic sucrose-density-gradient centrifugation. Four myelin subfractions, three floating respectively on 0.55 M- (light-myelin fraction), 0.75 M- (heavy-myelin fraction) and 0.85 M-sucrose (membrane fraction), and a pellet, were isolated and purified. At all ages, 70--75% of the total myelin proteins was found in the heavy-myelin fraction, whereas 2--5% of the protein was recovered in the light-myelin fraction, and about 7--12% in the membrane fraction. Most of the galactosyltransferase was associated with the heavy-myelin and membrane fractions. Other lipid-synthesizing enzymes studied appeared not to associate with purified myelin or myelin subfractions, but were enriched in the microsomal-membrane fraction. During development, the specific activity of the microsomal galactosyltransferase reached a maximum when the animals were about 20 days old and then declined. By contrast the specific activity of the galactosyltransferase in the heavy-myelin and membrane fractions was 3--4 times higher than that of the microsomal membranes in 16-day-old animals. The specific activity of the enzyme in the heavy-myelin fraction sharply declined with age. Chemical and enzymic analyses of the heavy-myelin and membrane myelin subfractions at various ages showed that the membrane fraction contained more proteins in relation to lipids than the heavy-myelin fraction. The membrane fraction was also enriched in phospholipids compared with cholesterol and contrined equivalent amounts of 2':3'-cyclic nucleotide 3'-phosphohydrolase compared with heavy- and light-myelin fractions. The membrane fraction was deficient in myelin basic protein and proteolipid protein and enriched in high-molecular-weight proteins. The specific localization of galactosyltransferase in heavy-myelin and membrane fractions at an early age when myelination is just beginning suggests that it may have some role in the myelination process.

Dataset Information

Molecular cloning of pigeon UDP-galactose:beta-D-galactoside alpha1,4-galactosyltransferase and UDP-galactose:beta-D-galactoside beta1,4-galactosyltransferase, two novel enzymes catalyzing the formation of Gal alpha1-4Gal beta1-4Gal beta1-4GlcNAc sequence.

Publications

Molecular cloning of pigeon UDP-galactose:beta-D-galactoside alpha1,4-galactosyltransferase and UDP-galactose:beta-D-galactoside beta1,4-galactosyltransferase, two novel enzymes catalyzing the formation of Gal alpha1-4Gal beta1-4Gal beta1-4GlcNAc sequence.

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