Research Group Bioanalytical Mass Spectrometry

Research Projects

We have a long standing interest in the analysis of protein-RNA interaction sites in UV irradiated ribonucleoprotein (RNP) particles. Over the last years we developed several strategies for the efficient analysis of cross-linking experiments. These include enrichment strategies making use of titanium dioxide, the employing photoreactive base-analogues for the enhancement of cross-linking yields as well as a novel data analysis strategy.

Protein-nucleic acid crosslinking

We have a long standing interest in the analysis of protein-RNA interaction sites in UV irradiated ribonucleoprotein (RNP) particles. Over the last years we developed several strategies for the efficient analysis of cross-linking experiments. These include enrichment strategies making use of titanium dioxide, the employing photoreactive base-analogues for the enhancement of cross-linking yields as well as a novel data analysis strategy.

Besides classical proteomics approaches that identify and quantifiy proteins, we are interested in how proteins directly interact with each other in complexes or signaling cascades. To address this question, we use cross-linking reagents that covalently connect individual protein binding partners and analyze the resulting cross-linked peptides by mass spectrometry. This enables the identification of protein interaction partners and furthermore provides structural information about protein folding and spatial interaction.

Protein-protein cross-linking

Besides classical proteomics approaches that identify and quantifiy proteins, we are interested in how proteins directly interact with each other in complexes or signaling cascades. To address this question, we use cross-linking reagents that covalently connect individual protein binding partners and analyze the resulting cross-linked peptides by mass spectrometry. This enables the identification of protein interaction partners and furthermore provides structural information about protein folding and spatial interaction.

SWATH (Selected Window Acquisition of all Theoretical precursors) analysis has emerged as a versatile approach to quantitatively profile whole proteome changes without the need for isotopic labeling. As a Data-Independent Acquisition (DIA) technique, it is particularly suited for rapid, parallelized analysis of complex biological samples. In conjunction with our research group and core facility at the University Medical Center Göttingen, we have established SWATH analysis and successfully applied it to a range of scientific projects, ranging from whole proteome characterization of bacteria and archaebacterial under environmental stresses to the analysis of tissue samples in biomedical research. 

Global label-free proteome profiling by SWATH analysis

SWATH (Selected Window Acquisition of all Theoretical precursors) analysis has emerged as a versatile approach to quantitatively profile whole proteome changes without the need for isotopic labeling. As a Data-Independent Acquisition (DIA) technique, it is particularly suited for rapid, parallelized analysis of complex biological samples. In conjunction with our research group and core facility at the University Medical Center Göttingen, we have established SWATH analysis and successfully applied it to a range of scientific projects, ranging from whole proteome characterization of bacteria and archaebacterial under environmental stresses to the analysis of tissue samples in biomedical research. 

In collaboration with the Department of Molecular Neurobiology (Prof. Reinhard Jahn) we investigate the proteome of the different building blocks of the synapse. We performed quantitative proteomic analyses of the active zone and synaptic vesicles protein inventory and analyzed protein phosphorylation in the presynapse. 

The synaptic proteome

In collaboration with the Department of Molecular Neurobiology (Prof. Reinhard Jahn) we investigate the proteome of the different building blocks of the synapse. We performed quantitative proteomic analyses of the active zone and synaptic vesicles protein inventory and analyzed protein phosphorylation in the presynapse. 

One source for protein diversity are post-translational protein modifications like phosphorylation, glycosylation, acetylation, methylation, ubiquitinylation, and SUMOylation. In collaboration with several research groups we apply methods for the enrichment and detection of post-translationally modified peptides by state-of-the-art mass spectrometry to investigate a multitude of biological questions.

Post-translational modifications

One source for protein diversity are post-translational protein modifications like phosphorylation, glycosylation, acetylation, methylation, ubiquitinylation, and SUMOylation. In collaboration with several research groups we apply methods for the enrichment and detection of post-translationally modified peptides by state-of-the-art mass spectrometry to investigate a multitude of biological questions.

One of the main building blocks of chromatin – the histone proteins, are subject to wide array of post-translational modifications. A major role of these marks is to recruit various protein factors which can bring about specific functional effects. In a close collaboration with the Guest Research Group Chromatin Biochemistry (Wolfgang Fischle) we aim at defining the complement of factors that is necessary and sufficient to translate single and patterns of chromatin marks into function. In a first step, we are using recombinant, defined chromatin templates to ‘fish’ for such factors using a pulldown strategy from cellular extracts in combination with quantitative mass spectrometry (SILAC). In a second step we are using bioinformatics approaches to systemically isolate chromatin interaction networks and to narrow down the list of chromatin factors essentially defining a chromatin domain.
 

 

Read-out of chromatin modification patterns

One of the main building blocks of chromatin – the histone proteins, are subject to wide array of post-translational modifications. A major role of these marks is to recruit various protein factors which can bring about specific functional effects. In a close collaboration with the Guest Research Group Chromatin Biochemistry (Wolfgang Fischle) we aim at defining the complement of factors that is necessary and sufficient to translate single and patterns of chromatin marks into function. In a first step, we are using recombinant, defined chromatin templates to ‘fish’ for such factors using a pulldown strategy from cellular extracts in combination with quantitative mass spectrometry (SILAC). In a second step we are using bioinformatics approaches to systemically isolate chromatin interaction networks and to narrow down the list of chromatin factors essentially defining a chromatin domain.

 
 
The exchange of macromolecules between the cytoplasm and the nucleus is mediated at the nuclear pore complex (NPC). This is a very large structure (~120 MDa in vertebrates) that spans inner and outer nuclear membranes. In vertebrates, it is composed of approximately 30 different nucleoporins (Nups) and it is estimated that the NPC contains approximately 600 individual proteins. In a collaboration with the Department of Molecular Biology at the University Medical Center Göttingen (Prof. Ralph Kehlenbach) and the Department of Cellular Logistics (Prof. Dirk Görlich) we investigate nucleocytoplasmic transport by state-of-the-art mass spectrometry.

Nucleocytoplasmic transport

The exchange of macromolecules between the cytoplasm and the nucleus is mediated at the nuclear pore complex (NPC). This is a very large structure (~120 MDa in vertebrates) that spans inner and outer nuclear membranes. In vertebrates, it is composed of approximately 30 different nucleoporins (Nups) and it is estimated that the NPC contains approximately 600 individual proteins. In a collaboration with the Department of Molecular Biology at the University Medical Center Göttingen (Prof. Ralph Kehlenbach) and the Department of Cellular Logistics (Prof. Dirk Görlich) we investigate nucleocytoplasmic transport by state-of-the-art mass spectrometry.

 
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