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Lysosomal storage diseases (LSD) are a group of ~ 50 inherited disorders with a cumulative incidence of ~ 1 in 5.000 new-borns. A prominent subgroup of LSDs originates from defects in sphingolipid degrading enzymes resulting in sphingolipid storage diseases such as Gaucher disease, Fabry disease or metachromatic leukodystrophy (MLD). Despite many years of research, the effects of these disorders on the cellular level are still poorly understood. To get a better insight on the processes involved, our group is investigating changes of the protein composition of cells as a consequence of increasing sphingolipid storage.

For this purpose, different animal and cell culture models of lipid storage diseases are analyzed using state of the art liquid chromatography tandem mass spectrometry methods.

We are using stable isotope based metabolic and chemical labeling strategies (e.g. stable isotope labeling of amino acids in cell culture (SILAC) [1] and tandem mass tags (TMT) [2]) in order to simoultaneously identify and quantify proteins and their changes on different levels of sample complexity up to whole cell proteomes.

We are also analysing posttranslational modifications and their quantitative patterns in the course of the disease. For the analysis of whole cell proteomes, multidimensional protein and peptide separation methods (Offgel electrophoresis and ion exchange chromatography [3]) are used to increase sample coverage which allows to identify tens of thousands of peptides and to quantify several thousand proteins from a single sample.

Using specific proteases, proteins are digested in peptides fractionated in subfractions and seperated using reversed phase chromatography followed by electrospray ionization and analysis using different state of the art mass spectrometers. Instruments available at the institute are 2 ion trap mass spectrometers, one LTQ-Orbitrap Velos mass spectrometer (a high performance hybrid tandem instrument) and a matrix assisted laser desorption ionization (MALDI) based instrument.


[1] Ong S.E., Blagoev B., Kratchmarova I., Kristensen D.B., Steen H., Pandey A., Mann M., Stable isotope labeling by amino acids in cell culture, SILAC, as a simple and accurate approach to expression proteomics. Mol Cell Proteomics. 2002 May;1(5):376-86.

[2] Thompson A., Schäfer J., Kuhn K., Kienle S., Schwarz J., Schmidt G., Neumann T., Johnstone R., Mohammed A.K., Hamon C., Tandem mass tags: a novel quantification strategy for comparative analysis of complex protein mixtures by MS/MS. Anal Chem. 2003 Apr 15;75(8):1895-904.

[3] Manadas, B., Mendes, V. M., English, J., Dunn, M. J., Peptide fractionation in proteomics approaches. Expert Rev. Proteomics 2010, 7, 655–663.


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