The Thermo Fisher Scientific Proteomics Facility for Disease Target Discovery

When you’re interested in knowing how protein abundances are regulated between different conditions. This can be done with a whole cell lysate, biofluids (e.g. plasma), or after enrichment of different sub-proteomes (e.g. mitochondrial purification).

Our facility uses mass spectrometry-based proteomics approaches to study PTMs in a comprehensive, unbiased manner. We have developed platforms for proteome-wide quantification of changes in many PTMs. Phosphorylation and ubiquitylation are the most commonly characterized PTMs. Less common are lysine acetylation or phosphotyrosine.

We use a variety of different techniques to study PPIs including:

Affinity-Purification mass spectrometry (AP-MS). This requires an affinity tagged bait (3xFLAG, Strep, GFP, etc.).

Endogenous protein immunoprecipitation for when you’re only interested in identifying or quantifying interaction partners for a single protein and we have a good antibody for that protein.

Co-elution mass spectrometry in which native lysates are separated by size-exclusion chromatography and complexes deduced by co-elution.

Proximity-Labeling mass spectrometry, which relies on fusion of a labeling enzyme (APEX, BirA, etc.) to the protein of interest. This method labels everything in ~10-20nm proximity of your fusion protein, but direct binding proteins, as well as those only in spatial proximity. Can be better for identifying transient interaction partners, at the cost of decreased specificity.

Methods utilized here include cross-linking mass spectrometry (XL-MS) to support integrative structural modeling, hydrogen-deuterium exchange mass spectrometry (HDX-MS) to characterize solvent accessibility, and native mass spectrometry to deduce protein mass and/or protein complex stoichiometry.