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Molecular & Cellular Proteomics 8:1708-1718, 2009.
© 2009 by The American Society for Biochemistry and Molecular Biology, Inc.

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Research

Mass Spectrometry-based Protein Profiling to Determine the Cause of Lysosomal Storage Diseases of Unknown Etiology^*,

David E. Sleat^,,¶, Lin Ding^,, Shudan Wang^||, Caifeng Zhao, Yanhong Wang, Winnie Xin^**, Haiyan Zheng, Dirk F. Moore, Katherine B. Sims^** and Katherine B. Sims^,,

From the Center for Advanced Biotechnology and Medicine and
Department of Pharmacology, University of Medicine and Dentistry of New Jersey-Robert Wood Johnson Medical School and
Department of Biostatistics, University of Medicine and Dentistry of New Jersey-School of Public Health, Piscataway, New Jersey 08854,
||McGill University, Montreal, Quebec H3A 2T5, Canada, and
**Neurology Department, Massachusetts General Hospital, Boston, Massachusetts 02114

Diagnosis of lysosomal storage diseases (LSDs) can be problematic in atypical cases where clinical phenotype may overlap with other genetically distinct disorders. In addition, LSDs may result from mutations in genes not yet implicated in disease. Thus, there are individuals that are diagnosed with apparent LSD based upon clinical criteria where the gene defect remains elusive. The objective of this study was to determine whether comparative proteomics approaches could provide useful insights into such cases. Most LSDs arise from mutations in genes encoding lysosomal proteins that contain mannose 6-phosphate, a carbohydrate modification that acts as a signal for intracellular targeting to the lysosome. We purified mannose 6-phosphorylated proteins by affinity chromatography and estimated relative abundance of individual proteins in the mixture by spectral counting of peptides detected by tandem mass spectrometry. Our rationale was that proteins that are decreased or absent in patients compared with controls could represent candidates for the primary defect, directing biochemical or genetics studies. On a survey of brain autopsy specimens from 23 patients with either confirmed or possible lysosomal disease, this approach identified or validated the genetic basis for disease in eight cases. These results indicate that this protein expression approach is useful for identifying defects in cases of undiagnosed lysosomal disease, and we demonstrated that it can be used with more accessible patient samples, e.g. cultured cells. Furthermore this approach was instrumental in the identification or validation of mutations in two lysosomal proteins, CLN5 and sulfamidase, in the adult form of neuronal ceroid lipofuscinosis.

To whom correspondence may be addressed: Center for Advanced Biotechnology and Medicine, 679 Hoes Lane, Piscataway, NJ 08854. Tel.: 732-235-5028; Fax: 732-235-4466; E-mail: lobel{at}cabm.rutgers.edu

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