Small Molecule Probes for Chemical Biology

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This proposal outlines a diverse program to further our work in chemical biology focusing on the development of tools to manipulate and study bacteria. We aim to develop a series of orthogonal probes that will allow monitoring of polysaccharide, protein, DNA and peptidoglycan synthesis, simultaneously, using imaging mass cytometry. On the path to these probes, new bioorthogonal chemistry, enzyme inhibitors and biological uses of tellurium will be developed. *** Through the synthesis of unnatural monosaccharides, which are recognized like their natural counterparts, we will produced compounds that target specific metabolic pathways. We hypothesize that through the judicious choice of the pathways targeted, it will be possible to label the polysaccharides produced by selected bacteria. We will develop new bioorthogonal chemistry to label these modified polysaccharides for detection. *** We aim to follow bacterial protein production with a novel phenylalanine analogue, containing a tellurophene, which is taken up and incorporated during protein synthesis. When used for monitoring global protein synthesis, the incorporation of tellurium can be followed by a recently developed atomic mass spectrometry technique called mass cytometry. The tellurophene also provides a useful tool for the analysis of recombinant proteins, the tellurium centre will provide a heavy atom for X-ray crystallography and an NMR active nucleus. The effect of incorporation of the modified amino acid in recombinant proteins will be studied. In addition, novel bioorthogonal chemistry at the tellurophene will be explored. *** Using imaging mass cytometry, we will apply our new polysaccharide and protein synthesis probes to answer questions about how bacteria form multicellular communities known as biofilms. Through employing a series of different tags it will be possible to monitor where and when polysaccharides, peptidoglycan, DNA and proteins are being synthesized. This highly parameterized assay will be the first example of how imaging mass cytometry can be coupled with multiple bioorthogonal chemistries to understand complex biological events. *** Overall this proposal encompasses substantial scientific breadth involving synthesis, enzymology, biophysical studies and bacterial assays. This research will provide an outstanding training ground for students to integrate their skills and solve problems at the interface of chemistry and biology. **