Membrane Mass Spectrometry
Project Group of Department of Molecular Membrane Biology (MMB)
The “Proteomics and membrane mass spectrometry” lab is funded by the MPIs for Biophysics and Brain Research. We specialize in the development of workflows, methods and – in collaboration with instrument vendors – mass spectrometers in order to address open questions in structural, molecular and neuro-biology. Our methods include:
- Direct, digest free sequencing of membrane protein complexes
- Single-crystal mass spectrometry
- Functional assays on membrane protein complexes
- Hydrogen-Deuterium-eXchange mass spectrometry (HDX-MS)
- Shotgun proteomics of complex biological samples
- Targeted proteomics of sub-proteomes (newly-synthesized and cell-type specific)
- Proteome turnover studies
- Post-Translational modifications
- MS Imaging
We operate eight instruments from Bruker, Thermo and Waters, including Orbitrap Fusion Lumos, Q Exactive plus, Impact-II, rapifleX, Synapt G2-Si HDMS, Orbitrap Elite, maXis and Autoflex III mass spectrometers. The systems are coupled to nano-UPLCs (Dionex U3000, easy-1200) and analytical UPLCs (U3000 and Acquity).
We also engage in external collaborations and are currently a member of SPP 2002 (Small proteins in prokaryotes) program by the DFG and the MOEL-SOEL-program by the BMBF.
Recent research highlights
Direct sequencing to identify novel subunits of membrane protein complexes.
Membrane proteins represent challenging targets for bottom-up proteomics. In particular small proteins lack proteolytic cleavage sites or don’t ionize efficiently in ESI. Together with Bruker Daltonics, we’ve modified a rapifleX MALDI-TOF/TOF mass spectrometer that now allows acquisition of information-rich PSD MS/MS spectra up to m/z 9000. We’ve successfully used this technique to identify multiple new subunits in membrane protein complexes that could not be identified previously (Kohlstädt et al., mBio, 2015; Safarian et al., Science, 2016; Bausewein et al., Cell, 2017).
HDX-MS of membrane protein complexes.
We successfully characterized ligand binding sites and activity-associated conformational dynamics in multiple membrane protein complexes, including a sodium:proton antiporter (Eisinger et al., PNAS, 2017) and a MATE transporter (Eisinger et al., JMB, 2018). We currently extend this technique to large membrane protein complexes.
We successfully characterized the proteome remodeling in homeostatic synaptic plasticity in primary hippocampal neurons, making use of the BONCAT technology (Schanzenbächer et al., Neuron, 2016; Schanzenbächer et al., eLife, 2018). We also monitored proteome turnover in this system (Dörrbaum et al., eLife, 2018), and currently investigate its role in homeostatic scaling. We also established a workflow allowing analysis of cell-type specific proteomes from living animals, and investigated proteome dynamics in animals exposed to an enriched environment (Alvarez-Castelao et al., Nature Biotechnology, 2017). A detailed description of the method is available at Alvarez-Castelao et al., Nature Protocols, 2019.
We recently characterized the molecular components in Sepia chromatophores, that play a key role in camouflage. Using a combination of UPLC-UV-ESI-MS/MS, MS Imaging and direct infusion UHR-MS, we identified Xanthomatin in these chromatophores, a compound that had previously been described to play a role in dragonfly coloration (Reiter et al., Nature, 2018).