title: Disease-causing mutations within the lysosomal integral membrane protein type 2 (LIMP-2) reveal the nature of binding to its ligand beta-glucocerebrosidase.
abstract: Action myoclonus-renal failure syndrome (AMRF) is caused by mutations in the lysosomal integral membrane protein type 2 (LIMP-2/SCARB2). LIMP-2 was identified as a sorting receptor for beta-glucocerebrosidase (beta-GC), which is defective in Gaucher disease. To date, six AMRF-causing mutations have been described, including splice site, missense and nonsense mutations. All mutations investigated in this study lead to a retention of LIMP-2 in the endoplasmic reticulum (ER) but affect the binding to beta-GC differentially. From the three nonsense mutations, only the Q288X mutation was still able to bind to beta-GC as efficiently as compared with wild-type LIMP-2, whereas the W146SfsX16 and W178X mutations lost their beta-GC-binding capacity almost completely. The LIMP-2 segment 145-288, comprising the nonsense mutations, contains a highly conserved coiled-coil domain, which we suggest determines beta-GC binding. In fact, disruption of the helical arrangement and amphiphatic nature of the coiled-coil domain abolishes beta-GC binding, and a synthetic peptide comprising the coiled-coil domain of LIMP-2 displays pH-selective multimerization properties. In contrast to the reduced binding properties of the nonsense mutations, the only missense mutation (H363N) found in AMRF leads to increased binding of beta-GC to LIMP-2, indicating that this highly conserved histidine modifies the affinity of LIMP-2 to its ligand. With the present study, we demonstrate that disruption of the coiled-coil structure or AMRF disease-causing mutations abolish beta-GC binding, indicating the importance of an intact coiled-coil structure for the interaction of LIMP-2 and beta-GC.
authors.affiliation: Institut für Biochemie, Christian-Albrechts-Universität zu Kiel, Olshausenstrasse 40, D-24098 Kiel, Germany.
authors.forename: Judith
authors.initials: J
authors.lastname: Blanz
authors.forename: Johann
authors.initials: J
authors.lastname: Groth
authors.forename: Christina
authors.initials: C
authors.lastname: Zachos
authors.forename: Christina
authors.initials: C
authors.lastname: Wehling
authors.forename: Paul
authors.initials: P
authors.lastname: Saftig
authors.forename: Michael
authors.initials: M
authors.lastname: Schwake
mesh_terms: D000818:Animals; D018955:CD36 Antigens / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004195:Disease Models, Animal; D005962:Glucosylceramidase / Q000235:genetics* / Q000378:metabolism*; D006801:Humans; D008024:Ligands; D051907:Lysosomal Membrane Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D018345:Mice, Knockout; D009154:Mutation; D020191:Myoclonic Epilepsies, Progressive / Q000235:genetics* / Q000378:metabolism; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D051116:Receptors, Scavenger / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
chemical_list: D018955:CD36 Antigens; D008024:Ligands; D051907:Lysosomal Membrane Proteins; D051116:Receptors, Scavenger; C074299:SCARB2 protein, human; C489452:Scarb2 protein, mouse; D005962:Glucosylceramidase