Mass spectrometry-based chemoproteomics for target deconvolution of bioactive natural products

Natural products play an important role in phenotypic approaches to drug discovery. Recent years have seen the resurgence of these approaches, in particular due to their pre-selection for cell-active compounds and their ability to identify novel druggable nodes. Therefore, the identification of protein targets of natural products and the elucidation of their mechanism of action remain areas of high interest. Quantitative chemoproteomics has emerged for us as a principal strategy for generation of testable target hypotheses. It typically combines competition-based compound affinity chromatography from cell lysates with mass spectrometry-based protein identification and quantitation using e.g. isobaric labeling tags. These studies enable the identification of the full spectrum of cellular interactors of compounds under conditions approximating the disease-relevant in vivo situation, including the efficacy target(s) and also potential off-targets. However, for unbiased target deconvolution, the immobilization of compound on solid support is required at some stage during the workflow. While the introduction of a functionalized linker at a permissive site, as typically done for synthetic small molecules, is in principle also possible for natural products, it requires extensive knowledge of structure activity relationships which might not always be easily obtainable. Therefore, an extended toolbox of approaches has proven to be valuable, including site-nonselective compound immobilization on photocrosslinker beads. Several variants of chemoproteomics as applied to natural products target deconvolution and validation will be presented.