Better sensitivity and higher throughout for palaeoproteomics

My project

The latest generation of high-resolution tandem mass spectrometers can reliably and rapidly detect proteins in complex mixtures, even at the very limited quantities typical of ancient samples. Despite these developments, ancient protein analysis is still in its infancy. Protein extraction and preparation for MS analysis is the first critical phase affecting recovery success. Simplification of sample preparation by integration of different chemical treatments: reduction, alkylation and digestion, into single-steps already proved to improve recovery in the most recalcitrant ancient samples. The recent availability of new high-resolution mass spectrometric instruments offers increased sensitivity in the low attomoles range. The possibility to combine complementary fragmentation methods (HCD + ETD) maximises the possibility to improve peptide identification and analyze ancient protein modifications with unprecedented depth and accuracy. Confident use of these technologies is still limited to very few specialized research units, with direct connections with instrument suppliers.

As an alternative to bottom-up proteomics, the emergence of new ion dissociation methods continues to drive top-down proteomics by offering valuable alternatives to traditional slow-heating methods (e.g., collision-activated dissociation, CAD). Electron transfer dissociation (ETD) leverages electron-driven radical rearrangements to promote cleavage of N–Cα bonds between amino acid residues, preserving labile post-translational modifications (PTMs) and providing extensive sequence-informative fragmentation of peptides and proteins. The complementary of the different types of fragmentation methods significantly increases confidence in peptide and protein identification in particular for PTM analyses. Therefore, top-down analysis allows for the measurement of intact protein masses and provides information on post-translational modifications, as well as the protein sequence via fragmentation of the intact proteins in the mass spectrometer. Application of top-down proteomics to cultural heritage materials will remove many of the limits hampering analysis of this category of samples.