title: Purification and biochemical characterization of a lysosomal α-fucosidase from the deuterostomia Asterias rubens.
abstract: In vertebrates, mannose 6-phosphate receptors [MPR300 (Mr 300 kDa) and MPR46 (Mr 46 kDa)] are highly conserved transmembrane glycoproteins that mediate transport of lysosomal enzymes to lysosomes. Our studies have revealed the appearance of these putative receptors in invertebrates such as the molluscs and deuterostomes. Starfish tissue extracts contain several lysosomal enzyme activities and here we describe the affinity purification of α-fucosidase. The purified enzyme is a glycoprotein that exhibited a molecular mass of ∼56 kDa in SDS-PAGE under reducing conditions. It has also cross-reacted with an antiserum to the mollusc enzyme suggesting antigenic similarities among the two invertebrate enzymes. LC-MS/MS analysis of the proteolytic peptides of the purified enzyme in combination with de novo sequencing allowed us to do partial amino acid sequence determination of the enzyme. These data suggest that this invertebrate enzyme is homologous to the known mammalian enzyme. The purified enzyme exhibited a mannose 6-phosphate dependent interaction with the immobilized starfish MPR300 protein. Our results demonstrate that the lysosomal enzyme targeting pathway is conserved even among the invertebrates.
authors.affiliation: Protein Biochemistry and Molecular Biology Laboratory, Department of Biochemistry, University of Hyderabad, Hyderabad 500 046, India.
authors.forename: Merino
authors.initials: M
authors.lastname: Visa
authors.forename: Elke
authors.initials: E
authors.lastname: Hammer
authors.forename: Uwe
authors.initials: U
authors.lastname: Völker
authors.forename: Hendrik
authors.initials: H
authors.lastname: Koliwer-Brandl
authors.forename: Sørge
authors.initials: S
authors.lastname: Kelm
authors.forename: Siva Kumar
authors.initials: SK
authors.lastname: Nadimpalli
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D047271:Asterias / Q000201:enzymology*; D008247:Lysosomes / Q000201:enzymology*; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D017386:Sequence Homology, Amino Acid; D005644:alpha-L-Fucosidase / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism*
chemical_list: D005644:alpha-L-Fucosidase