Molecular Cloning and Characterization of Human GnT-IX, a Novel β1,6-N-Acetylglucosaminyltransferase That Is Specifically Expressed in the Brain

Abstract

A novel ␤1,6-N-acetylglucosaminyltransferase (␤1, 6GnT) cDNA was identified by a BLAST search using the amino acid sequence of human GnT-V as a query. The full-length sequence was determined by a combination of 5-rapid amplification of cDNA end analysis and a further data base search. The open reading frame encodes a 792 amino acid protein with a type II membrane protein structure typical of glycosyltransferases. The entire sequence identity to human GnT-V is 42%. When pyridylaminated (PA) agalacto biantennary N-linked oligosaccharide was used as an acceptor substrate, the recombinant enzyme generated a novel product other than the expected GnT-V product, (GlcNAc␤1,2-Man␣1,3-)[GlcNAc␤1,2-(GlcNAc␤1,6-)Man␣1,6-]Man␤1,4-GlcNAc␤1,4-GlcNAc-PA. This new product was identified as [GlcNAc␤1,2-(GlcNAc␤1,6-)Man␣1,3-][GlcNAc␤1,2-(GlcNAc␤1,6-)Man␣1,6-]Man␤1,4-GlcNAc␤1,4-GlcNAc-PA by mass spectrometry and 1 H NMR. Namely, the new GnT (designated as GnT-IX) has ␤1,6GnT activity not only to the ␣1,6-linked mannose arm but also to the ␣1,3-linked mannose arm of N-glycan, forming a unique structure that has not been reported to date. Northern blot analysis showed that the GnT-IX gene is exclusively expressed in the brain, whereas the GnT-V gene is expressed ubiquitously. These results suggest that GnT-IX is responsible for the synthesis of a unique oligosaccharide structure in the brain.

Human Prostate Cancer in a Clinically Relevant Xenograft Mouse Model: Identification of β(1,6)-Branched Oligosaccharides as a Marker of Tumor Progression

“…In contrast to previously used "leaky" SCID mice, pfp (32,33). Mgat5b has so far exclusively been described in neural tissues and testes, whereas Mgat5 is expressed in most human and rodent tissues (23,34). This work therefore describes increased expression levels of Mgat5b in human prostate cancer cells compared with primary human prostate epithelium for the first time (Fig.…”

A Route from Glycobiology to Congenital Muscular Dystrophy

“…A gene for such a 6-branching enzyme (GnT-IX, GnT-VB) was shown to be specifically expressed in the brain (29)(30)(31).…”

Biosynthetic Pathway of O-Mannosyl Glycan in Mammals

“…1). The GlcNAcβ1-6Man linkage was formed by GnT-IX and GnT-VB that were specifically expressed in brain (14)(15)(16). Because the transfer of GlcNAc to the C6-position of O-mannose by GnT-IX or GnT-VB required the GlcNAcβ1-2Man structure, POMGnT1 must work before GnT-IX or GnT-VB in O-Man glycan biosynthesis (Fig.…”

“…1), such as HNK-1, SSEA-1, GlcNAcβ1-2(GlcNAcβ1-6)Man, GalNAcβ1-3GlcNAcβ1-4Man and GalNAcβ1-3GlcNAcβ1-4Man-6-phosphodiester (14)(15)(16)(17). Although it is still unknown what kind of structure attaches to the phosphodiester linkage of O-mannose, the possibility that the phosphodiester linkage is required for the laminin-binding activity of α-DG was reported (17).…”