Mass spectrometry-based protein profiling to determine the cause of lysosomal storage diseases of unknown etiology.
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.
Sleat
David E
DE
Center for Advanced Biotechnology and Medicine, Department of Pharmacology, University of Medicine and Dentistry of New Jersey-Robert Wood Johnson Medical School, Piscataway, New Jersey 08854, USA.
Ding
Lin
L
Wang
Shudan
S
Zhao
Caifeng
C
Wang
Yanhong
Y
Xin
Winnie
W
Zheng
Haiyan
H
Moore
Dirk F
DF
Sims
Katherine B
KB
Lobel
Peter
P
D000328:Adult; D000595:Amino Acid Sequence; D001923:Brain Chemistry; D002648:Child; D016000:Cluster Analysis; D005260:Female; D006023:Glycoproteins / Q000737:chemistry / Q000302:isolation & purification; D006801:Humans; D007223:Infant; D016464:Lysosomal Storage Diseases / Q000209:etiology / Q000235:genetics / Q000378:metabolism; D008297:Male; D008360:Mannosephosphates / Q000737:chemistry; D013058:Mass Spectrometry / Q000379:methods*; D008969:Molecular Sequence Data; D010802:Phylogeny; D040081:Protein Array Analysis; D011506:Proteins / Q000737:chemistry / Q000235:genetics; D020543:Proteome / Q000032:analysis*; D040901:Proteomics / Q000379:methods; D053719:Tandem Mass Spectrometry / Q000379:methods; D055815:Young Adult
D006023:Glycoproteins; D008360:Mannosephosphates; D011506:Proteins; D020543:Proteome; C018589:lysosomal proteins; C027693:mannose-6-phosphate