title: Biochemical and immunological characterization of a glycosylated alpha-fucosidase from the invertebrate Unio: interaction of the enzyme with its in vivo binding partners.
abstract: Mammalian alpha-fucosidase (EC 3.2.1.51) is a lysosomal enzyme that catalyzes the removal of fucose residues from glycosphingolipids and its absence in humans results in a rare metabolic disorder called fucosidosis. Among the invertebrates in the molluscs (Unio) two forms of the enzyme have been reported, a 68 kDa non-glycosylated form and a 56 kDa glycosylated form. The glycosylated form has been purified from the seminal fluid of Unio [Biochem. Biophys. Res. Commun. 234 (1997) 54]. In the present study, the 56 kDa glycosylated form has been purified to homogeneity from the whole body tissue of Unio using a series of chromatographic steps. The purified enzyme migrated as a single protein species in 10% SDS-PAGE. Antibodies to the purified enzyme were raised in a rabbit in order to study its biochemical and immunological properties. The purified enzyme is a glycoprotein that exhibits strong binding to Con A-Sepharose gel and can be deglycosylated by PNGase F enzyme suggesting it to be N-glycosylated. The enzyme has been shown to specifically interact with the mannose 6-phosphate receptor protein (MPR 300) purified from goat and Unio. This specific interaction is discussed in view of its possible in vivo binding partners.
authors.affiliation: Laboratory of Protein Biochemistry and Molecular Biology, Department of Biochemistry, University of Hyderabad, Hyderabad 500 046, India. drnsk7@yahoo.co.in
authors.forename: Siva Kumar
authors.initials: SK
authors.lastname: Nadimpalli
authors.forename: Nirmala
authors.initials: N
authors.lastname: Padmanabhan
authors.forename: Suresh
authors.initials: S
authors.lastname: Koduru
mesh_terms: D000818:Animals; D002845:Chromatography / Q000379:methods; D003208:Concanavalin A / Q000737:chemistry; D004591:Electrophoresis, Polyacrylamide Gel; D006023:Glycoproteins / Q000737:chemistry; D006031:Glycosylation; D006801:Humans; D008024:Ligands; D008247:Lysosomes / Q000737:chemistry; D008974:Mollusca; D043524:Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase / Q000737:chemistry; D011485:Protein Binding; D012685:Sepharose / Q000737:chemistry; D005644:alpha-L-Fucosidase / Q000737:chemistry* / Q000276:immunology / Q000302:isolation & purification
chemical_list: D006023:Glycoproteins; D008024:Ligands; D003208:Concanavalin A; D012685:Sepharose; D005644:alpha-L-Fucosidase; D043524:Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase