Expression and characterization of 14 GLB1 mutant alleles found in GM1-gangliosidosis and Morquio B patients

Raül Santamaria^*^,^,, Amparo Chabás^,**, John W. Callahan, Daniel Grinberg¹^,*^,^, and Lluïsa Vilageliu¹^,2^,*^,^,

^* Departament de Genètica, Universitat de Barcelona, Barcelona, Spain
Centero de Investigación Biomédica en red de Enfermedades Raras, Barcelona Spain
Institut de Biomedicina de la Universitat de Barcelona, Barcelona Spain
^** Institut de Bioquímica Clínica, Hospital Clínic, Corporació Sanitària Clínic, Barcelona, Spain
Research Institute, Hospital of Sick Children, Toronto, Canada

Published, JLR Papers in Press, July 30, 2007.

^¹ D. Grinberg and L. Vilageliu are co-last authors.

^² To whom correspondence should be addressed. e-mail: lvilageliu{at}ub.edu

GM1-gangliosidosis and Morquio B disease are lysosomal storagedisorders caused by ß-galactosidase deficiency attributableto mutations in the GLB1 gene. On reaching the endosomal-lysosomalcompartment, the ß-galactosidase protein associateswith the protective protein/cathepsin A (PPCA) and neuraminidaseproteins to form the lysosomal multienzyme complex (LMC). Thecorrect interaction of these proteins in the complex is essentialfor their activity. More than 100 mutations have been describedin GM1-gangliosidosis and Morquio B patients, but few have beenfurther characterized. We expressed 12 mutations suspected tobe pathogenic, one known polymorphic change (p.S532G), and avariant described as either a pathogenic or a polymorphic change(p.R521C). Ten of them had not been expressed before. The expressionanalysis confirmed the pathogenicity of the 12 mutations, whereasthe relatively high activity of p.S532G is consistent with itsdefinition as a polymorphism. The results for p.R521C suggestthat this change is a low-penetrant disease-causing allele.Furthermore, the effect of these ß-galactosidase changeson the LMC was also studied by coimmunoprecipitations and Westernblotting. The alteration of neuraminidase and PPCA patternsin several of the Western blotting analyses performed on patientprotein extracts indicated that the LMC is affected in at leastsome GM1-gangliosidosis and Morquio B patients.

Supplementary key words ß-galactosidase • heterologous expression • lysosomal multienzyme complex

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