Assessment of glycosaminoglycan-protein linkage tetrasaccharides as acceptors for GalNAc- and GlcNAc-transferases from mouse mastocytoma.

Two glycosaminoglycan-protein linkage tetrasaccharide-serine compounds, GlcAbeta1-3Galbeta1-3Galbeta1-4Xylbeta1-O-Ser and GlcAbeta1-3Gal(4-O-sulfate)beta1-3Galbeta1-4Xylbeta1-O -Ser, were tested as hexosamine acceptors, using UDP-[3H]GlcNAc and UDP-[3H]GalNAc as sugar donors, and solubilized mouse mastocytoma microsomes as enzyme source. The nonsulfated Ser-tetrasaccharide was found to function as an acceptor for a GalNAc residue, whereas the Ser-tetrasaccharide containing a sulfated galactose unit was inactive. Characterization of the radio-labelled product by digestion with alpha-N-acetylgalactosaminidase and beta-N-acetylhexosaminidase revealed that the [3H]GalNAc unit was alpha-linked, as in the product previously synthesized using serum enzymes, and not beta-linked as found in the chondroitin sulfate polymer.

Biosynthesis of the escherichia coli K4 capsule polysaccharide. A parallel system for studies of glycosyltransferases in chondroitin formation.

Escherichia coli K4 bacteria synthesize a capsule polysaccharide (GalNAc-GlcA(fructose))n with the carbohydrate backbone identical to chondroitin. GlcA- and GalNAc-transferase activities from the bacterial membrane were assayed with acceptors derived from the capsule polysaccharide and radiolabeled UDP-[14C]GlcA and UDP-[3H]GalNAc, respectively. It was shown that defructosylated oligosaccharides (chondroitin) could serve as substrates for both the GlcA- and the GalNAc-transferases.

Substrate specificities of glycosyltransferases involved in formation of heparin precursor and E. coli K5 capsular polysaccharides.

The E. coli K5 capsular polysaccharide is composed of 4)GlcpA(beta 1-4)GlcpNAc(alpha 1-disaccharide units. A partially N-deacetylated/N-sulfated heptasaccharide, derived from this polymer and having a nonreducing terminal GlcNAc unit, was used as acceptor for a mastocytoma microsomal GlcA-transferase involved in heparin biosynthesis.