Sugar-binding sites with specificity to N-acetyl-D-glucosaminides in middle ear mucosa of the guinea pig.

Abstract

The distribution of sugar-binding sites was analyzed in Lowicryl K4M-embedded guinea pig middle ear mucosa. Four neoglycoproteins and a glycoprotein were used as probes: N-acetyl-D-glucosamine (GlcNAc), D-mannose, N-acetyl-D-galactosamine, or L-fucose carrying bovine serum albumin (BSA) and asialofetuin with terminal D-galactosyl sugar residues. Each probe was then labelled with 15 nm colloidal gold. Incubation of ultrathin sections with gold-labelled p-aminophenyl N-acetyl-beta-D-glucosaminide-BSA (GlcNAc/BSA/gold) led to binding on mucosal cilial, microvilli, rough endoplasmic reticulum, mitochondria, and nuclei. No binding occurred with asialofetuin or neoglycoproteins containing mannose, N-acetylgalactosamine or fucose. Various control experiments showed that specificity of GlcNAc/BSA/gold binding was directed towards N-acetylglucosaminyl residues expressed on the neoglycoprotein. Competitive sugar inhibition with GlcNAc and its derivatives suggested that the strong affinity for GlcNAc-binding sites took place in a complex formation with sugar residues bound to a carrier protein. The existence of a hydrophobic region close to the sugar-binding site was also suggested.