Molecular cloning of a novel human UDP-GalNAc:polypeptide N-acetylgalactosaminyltransferase, GalNAc-T8, and analysis as a candidate autosomal dominant hypophosphatemic rickets (ADHR) gene

173

148

The completed fruit fly genome was found to contain up to 15 putative UDP-N-acetyl-␣-D-galactosamine:polypeptide N-acetylgalactosaminyltransferase (GalNAc-transferase) genes. Phylogenetic analysis of the putative catalytic domains of the large GalNAc-transferase enzyme families of Drosophila melanogaster (13 available), Caenorhabditis elegans (9 genes), and mammals (12 genes) indicated that distinct subfamilies of orthologous genes are conserved in each species. In support of this hypothesis, we provide evidence that distinctive functional properties of Drosophila and human GalNAc-transferase isoforms were exhibited by evolutionarily conserved members of two subfamilies (dGalNAc-T1 (l(2)35Aa) and GalNAc-T11; dGalNAc-T2 (CG6394) and GalNAc-T7). dGalNAc-T1 and novel human GalNAc-T11 were shown to encode functional GalNActransferases with the same polypeptide acceptor substrate specificity, and dGalNAc-T2 was shown to encode a GalNAc-transferase with similar GalNAc glycopeptide substrate specificity as GalNAc-T7. Previous data suggested that the putative GalNAc-transferase encoded by l(2)35Aa had a lethal phenotype (Flores, C., and Engels, W. (1999) Proc. Natl. Acad. Sci. U. S. A. 96, 2964-2969), and this was substantiated by sequencing of three lethal alleles l(2)35Aa HG8

Section: Galnac-transferase Subfamily Dgalnac-t1(l(2)35aa)/human Galnmentioning

confidence: 99%

Functional Conservation of Subfamilies of Putative UDP-N-acetylgalactosamine:Polypeptide N-Acetylgalactosaminyltransferases inDrosophila, Caenorhabditis elegans, and Mammals

Schwientek¹,

Bennett²,

Flores³

et al. 2002

173

148

“…These results show that GalNAc-T4 can glycosylate Ser 20 in -GSTA-efficiently, in addition to the two sites previously identified (7), provided the substrate has GalNAc residues at Thr 9 and/or Thr 21 . The kinetics of the reaction is such that it is not possible to identify which of the two sites, Thr 14 or Ser 20 , is first glyco- (13)(14)(15)(16)(17)(18)(19)(20)(21)(22)(23)(24)(25)ϩ3GalNAc. The identified degradation products are labeled in the spectra according to TAP25 peptide sequence.…”

Section: Galnac-t4 Transfer To At Least Three Sites In the Muc1mentioning

confidence: 99%

“…[1][2][3]. To date seven members of the mammalian GalNAc-transferase family have been identified and characterized (4 -12), and several additional putative members of this gene family have been predicted from analysis of genome data bases (13). The GalNAc-transferase isoforms have different kinetic properties and show differential expression patterns temporally and spatially, suggesting that they have distinct biological functions (2).…”

mentioning

confidence: 99%

The Lectin Domain of UDP-N-acetyl-d-galactosamine:PolypeptideN-acetylgalactosaminyltransferase-T4 Directs Its Glycopeptide Specificities

Hassan¹,

Reis

Bennett³

et al. 2000

152

137

The initiation step of mucin-type O-glycosylation is controlled by a large family of homologous UDP-GalNAc: polypeptide N-acetylgalactosaminyltransferases (GalNAc-transferases). Differences in kinetic properties, substrate specificities, and expression patterns of these isoenzymes provide for differential regulation of O-glycan attachment sites and density. Recently, it has emerged that some GalNAc-transferase isoforms in vitro selectively function with partially GalNAc O-glycosylated acceptor peptides rather than with the corresponding unglycosylated peptides. O-Glycan attachment to selected sites, most notably two sites in the MUC1 tandem repeat, is entirely dependent on the glycosylation-dependent function of GalNAc-T4. Here we present data that a putative lectin domain found in the C terminus of GalNAc-T4 functions as a GalNAc lectin and confers its glycopeptide specificity. A single amino acid substitution in the lectin domain of a secreted form of GalNAc-T4 selectively blocked GalNAc-glycopeptide activity, while the general activity to peptides exerted by this enzyme was unaffected. Furthermore, the GalNAc-glycopeptide activity of wild-type secreted GalNAc-T4 was selectively inhibited by free GalNAc, while the activity with peptides was unaffected.

“…pp-GalNAc-Ts are biologically important because they determine the number and position of mucin-type O-glycans in a protein. To date, at least 11 human pp-GalNAc-Ts , -T1, -T2, -T3, -T4, -T6, -T7, -T8, -T9, -T10, -T11, and -T12, have been identified (5)(6)(7)(8)(9)(10)(11)(12)(13)(14)(15)(16). The proteins are 40 -60% identical in their sequence and are therefore homologous.…”

mentioning

confidence: 99%

Initiation of O-Glycan Synthesis in IgA1 Hinge Region Is Determined by a Single Enzyme, UDP-N-Acetyl-α-d-galactosamine:PolypeptideN-Acetylgalactosaminyltransferase 2

Iwasaki

Zhang

Tachibana

et al. 2003

The hinge region of human immunoglobulin A1 (*IgA1) possesses multiple O-glycans, of which synthesis is initiated by the addition of GalNAc to serine or threonine through the activity of UDP-N-acetyl-␣-D-galactosamine:polypeptide N-acetylgalactosaminyltransferases (pp-GalNAc-Ts). We found that six pp-GalNAc-Ts, ppGalNAc-T1, -T2, -T3, -T4, -T6, and -T9, were expressed in B cells, IgA-bearing B cells, and NCI-H929 IgA myeloma cells. pp-GalNAc-T activities of these six enzymes for a synthetic IgA hinge peptide, which has nine possible O-glycosylation sites, were examined using a reversed phase-high performance liquid chromatography, a matrix-assisted laser desorption ionization time of flight mass spectrometry, and peptide sequencing analysis. pp-GalNAc-T2 showed the strongest activity transferring GalNAc to a maximum of eight positions. Other pp-GalNAc-Ts exhibited different substrate specificities from pp-GalNAc-T2; however, their activities were extremely weak. It was reported that the IgA1 hinge region possesses a maximum of five O-glycans, and their amino acid positions have been determined. We found that pp-GalNAc-T2 selectively transferred GalNAc residues to the same five positions. These results strongly suggested that pp-GalNAc-T2 is an essential enzyme for initiation of O-linked glycosylation of the IgA1 hinge region.