Characterisation of a Novel Benzopyran Library Using High-Throughput Microscopy

Abstract

<p>Drug discovery is a multi-disciplinary field incorporating both chemistry and biology to create novel pharmaceuticals. Nature synthesizes a diverse range of chemical entities that can demonstrate a wide range of biological interactions, though often produces these compounds in small amounts. Using natures structural diversity as a template, organic synthetic chemistry can tap into the structures of natural products and provide novel structures as well as overcome supply issues through large-scale synthetic chemical processes. A novel benzopyran library was synthesised by Sandile Simelane by reacting 3,4,6,-tri-O-acetyl-D-galactal with various phenols to create a novel focused library of bridged benzopyrans. Each molecule has unique functional groups at defined points in the structure due to varying the functional groups on the phenol, allowing for variation within the library whilst retaining the core scaffold. In this thesis, the bioactivity of this novel benzopyran library was explored using a phenotypic screen measuring growth inhibition. A compound, S13, was determined to be the most potent in the library, therefore genome-wide screening was performed using S13. High-throughput microscopy of 4,100 strains, each with a different GFP-tagged protein, was utilized to determine proteins that increased in abundance or changed localization in response to perturbation with S13. Following treatment with S13, the yeast vacuole increased in size due to an aggregation of proteins in the vacuolar lumen. The increase in vacuole size was coincident with a decrease in vacuolar acidity, potentially disrupted autophagy and the upregulation of several proteins involved in ergosterol biosynthesis. Together, these results reveal a novel bridged benzopyran that increases vacuolar size and pH through an epistatic mechanism involving ergosterol biosynthesis.</p>