Chemical proteomics includes a powerful set of complementary, mass spectrometry based and proteome-wide approaches for identifying the key proteins relevant for producing the phenotypic effects of a treatment, including direct and indirect interactions with as well as mechanism of action. Chemical proteomics includes affinity capture approaches with: 1) compound-centric approaches using modified, tagged compounds as baits to identify target proteins that bind to them in a proteome-wide background; 2) activity based approaches, where irreversibly binding baits probe the activity status of broad pre-defined enzyme families. This method can elucidate target activity in vivo as well as in vitro in competitive binding assays. Another way to probe the effect of a chemical compound on the proteome is using thermal profiling. Thermal profiling proteomics exploits the fact that protein denaturation temperatures can be affected, both positively and negatively, when the system is perturbed with a chemical compound. Proteome-wide mapping of these thermal changes helps to understand the mechanisms of molecular changes induced in the cell by the chemical compound. Proteome-wide probing is performed by series of quantitative proteomics analyses of the purified soluble fraction after temperature-induced protein precipitation. To decipher the key proteins in the mechanism of action, quantitative proteomics can also be utilized by evaluating the specific regulation of certain proteins due to a certain treatment. This can be achieved using a specifically designed multiple drugs/compounds approach, also known as FITExP (functional identification of targets by expression proteomics). Moreover, the binding site of a chemical compound to a particular protein can be probed by deuterium exchange experiments using mass spectrometry.
Support within chemical proteomics at BioMS include:
- Affinity capture using click chemistry based probes followed by MS-analysis, in collaboration with national drug development and chemical biology infrastructures (who can help with the organic synthesis of needed affinity probes).
- Kinetic profiling post compound exposure
- Thermal profiling proteomics
- Specifically designed quantitative proteomics using the multiple drugs/compounds FITExP method
- Protein characterization by top-down proteomics
- Interaction interface elucidation by deuterium exchange mass spectrometry assays