title: Identification of the interaction domains between α- and γ-subunits of GlcNAc-1-phosphotransferase.
abstract: The disease-associated hexameric N-acetylglucosamine (GlcNAc)-1-phosphotransferase complex (α2β2γ2) catalyzes the formation of mannose 6-phosphate residues on lysosomal enzymes required for efficient targeting to lysosomes. Using pull-down experiments and mutant subunits, we identified a potential loop-like region in the α-subunits comprising residues 535-588 and 645-698 involved in the binding to γ-subunits. The interaction is independent of the mannose 6-phosphate receptor homology domain but requires the N-terminal unstructured part of the γ-subunit consisting of residues 26-69. These studies provide new insights into structural requirements for the assembly of the GlcNAc-1-phosphotransferase complex, and the functions of distinct domains of the α- and γ-subunits.
authors.affiliation: Section Biochemistry, Children's Hospital, University Medical Center Hamburg-Eppendorf, Germany.
authors.forename: Renata Voltolini
authors.initials: RV
authors.lastname: Velho
authors.affiliation: Section Biochemistry, Children's Hospital, University Medical Center Hamburg-Eppendorf, Germany.
authors.forename: Raffaella
authors.initials: R
authors.lastname: De Pace
authors.affiliation: Institute for Biochemistry and Molecular Biology, University of Hamburg, Germany.
authors.forename: Henning
authors.initials: H
authors.lastname: Tidow
authors.affiliation: Section Biochemistry, Children's Hospital, University Medical Center Hamburg-Eppendorf, Germany.
authors.forename: Thomas
authors.initials: T
authors.lastname: Braulke
authors.affiliation: Section Biochemistry, Children's Hospital, University Medical Center Hamburg-Eppendorf, Germany.
authors.forename: Sandra
authors.initials: S
authors.lastname: Pohl
mesh_terms: D000117:Acetylglucosamine / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D057809:HEK293 Cells; D006801:Humans; D009154:Mutation; D010641:Phenotype; D010770:Phosphotransferases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D000072417:Protein Domains; D021122:Protein Subunits / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*
chemical_list: D021122:Protein Subunits; D010770:Phosphotransferases; D000117:Acetylglucosamine
keywords: MRH domain; hexameric GlcNAc-1-phosphotransferase complex; lysosomal enzymes; mannose 6-phosphate; mucolipidosis II and III; pull-down interaction approach