Improved deconvolution of natural products' protein targets using diagnostic ions from chemical proteomics linkers

Identification of interactions between proteins and natural products or similar active small molecules is crucial for understanding of their mechanism of action on a molecular level. To search elusive, often labile, and low-abundant conjugates between proteins and active compounds, chemical proteomics introduces a feasible strategy that allows to enrich and detect these conjugates. Recent advances in mass spectrometry techniques and search algorithms provide unprecedented depth of proteome coverage and the possibility to detect desired modified peptides with high sensitivity. The chemical 'linker' connecting an active compound-protein conjugate with a detection tag is the critical component of all chemical proteomic workflows. In this review, we discuss the properties and applications of different chemical proteomics linkers with special focus on their fragmentation releasing diagnostic ions and how these may improve the confidence in identified active compound-peptide conjugates. The application of advanced search options improves the identification rates and may help to identify otherwise difficult to find interactions between active compounds and proteins, which may result from unperturbed conditions, and thus are of high physiological relevance.