Objective

Understanding how a bioactive molecule works is essential in many aspects of life sciences and specifically in the drug discovery and development cycle. Projects initiated via phenotypic screening approaches need target deconvolution methods to progress more efficiently.

We have developed ULTImate YChemH, a powerful chemical biology tool to identify the direct protein targets of small molecules. It is a unique screening platform based on an improved Chemical Yeast Three-Hybrid technique. Highly complex protein domain libraries, prepared from any cell type or tissue, are screened to saturation to identify partners and their interacting domains using a transcriptional read-out. Sophisticated bioinformatics analysis allows to attribute confidence scores to each interaction. This in-yeast screening technology has been optimized for small molecules with a special emphasis on chemical derivatization and the generation of permeable yeast strains. Our versatile ULTImate YChemH technique is complementary to other proteomics technologies, with several key advantages: it is an unbiased, in vivo screening technology as it screens the entire proteome from a given tissue or cell line. The absence of any washing steps contributes to its high sensitivity. In addition, each putative interaction partner is tested individually, eliminating the competition by abundant or strong binders.

With the overarching goal of developing new therapies for diabetes, an unbiased chemical-genetic screens in zebrafish was performed to identify compounds, signals and cellular mechanisms that promote beta-cell regeneration. Hit compounds were selected for optimization and targets deconvolution. Ultimate YChemH was used for identifying targets of the most potent hit compound. It resulted in a short-list of 3 top candidate targets, out of which one was confirmed to directly interact with the compound using surface plasmon resonance. A chemically divert inhibitor of the target protein displayed similar biological effects as our hit, adding confidence to the proposed target and the screening rationale.