Affinity matrices to profile HDAC inhibitors
Linkable analogues of molecules known to inhibit histone deacetylases are synthesised (organic chemistry) and immobilised on sepharose beads. The affinity matrices hence obtained are tested for their ability to enrich HDACs out of cell or tissue lysates. The best matrices are used to profile inhibitors for their binding to native state proteins (i.e. engulfed in complexes and containing PTMs): the inhibitors – in increasing concentrations – and the matrices are competing for binding in the active site of the HDACs present in the lysate; only the fraction bound by the matrices is enriched and subsequently quantified by bottom-up proteomics allowing to obtain inhibition curves for native proteins.