PROTEASE DEGRADOMICS STUDIED USING POSITIONAL PROTEOMICS

Positional proteomics refers to reductionist proteomics approaches by which "terminal peptides" - mainly amino (N) terminal, tough more recently also carboxy (C) terminal peptides - are enriched via (affinity) chromatography approaches, and then analyzed by contemporary mass spectrometry. Back in 2003, our lab was the first to introduce these approaches by means of a diagonal chromatography (COFRADIC) technology to enrich for N-terminal peptides in tryptic digests of proteomes of
human platelets.
Having access to such protein termini allows for the study of protein processing events mediated by proteases. I will discuss the current status, opportunities, challenges and caveats of this type of mass spectrometry driven research. More in particular, a novel technology to enrich for C-terminal peptides, with applications to degradomics of carboxypeptidases, will be introduced. Further, I will discuss technologies that allow straightforward distinction between efficient protease substrates and bystanders, the latter which are highly likely not physiologically relevant,
and I will explain how characterization of protease substrates can be automated, with a link to a novel methodology for screening protein-protein interactions.