Specific enzyme complex of beta-1,4-galactosyltransferase-II and glucuronyltransferase-P facilitates biosynthesis of N-linked human natural killer-1 (HNK-1) carbohydrate.

ABSTRACT: Human natural killer-1 (HNK-1) carbohydrate is highly expressed in the nervous system and is involved in synaptic plasticity and dendritic spine maturation. This unique carbohydrate, consisting of a sulfated trisaccharide (HSO(3)-3GlcA?1-3Gal?1-4GlcNAc-), is biosynthesized by the successive actions of ?-1,4-galactosyltransferase (?4GalT), glucuronyltransferase (GlcAT-P and GlcAT-S), and sulfotransferase (HNK-1ST). A previous study showed that mice lacking ?4GalT-II, one of seven ?4GalTs, exhibited a dramatic loss of HNK-1 expression in the brain, although ?4GalT-I-deficient mice did not. Here, we investigated the underlying molecular mechanism of the regulation of HNK-1 expression. First, focusing on a major HNK-1 carrier, neural cell adhesion molecule, we found that reduced expression of an N-linked HNK-1 carbohydrate caused by a deficiency of ?4GalT-II is not likely due to a general loss of the ?1,4-galactose residue as an acceptor for GlcAT-P. Instead, we demonstrated by co-immunoprecipitation and endoplasmic reticulum-retention analyses using Neuro2a (N2a) cells that ?4GalT-II physically and specifically associates with GlcAT-P. In addition, we revealed by pulldown assay that Golgi luminal domains of ?4GalT-II and GlcAT-P are sufficient for the complex to form. With an in vitro assay system, we produced the evidence that the kinetic efficiency k(cat)/K(m) of GlcAT-P in the presence of ?4GalT-II was increased about 2.5-fold compared with that in the absence of ?4GalT-II. Finally, we showed that co-expression of ?4GalT-II and GlcAT-P increased HNK-1 expression on various glycoproteins in N2a cells, including neural cell adhesion molecule. These results indicate that the specific enzyme complex of ?4GalT-II with GlcAT-P plays an important role in the biosynthesis of HNK-1 carbohydrate.

SUBMITTER: Kouno T 

PROVIDER: S-EPMC3173110 | biostudies-literature | 2011 Sep

REPOSITORIES: biostudies-literature