THESIS TITLE: Characterizing Surface Thiol Sites of Environmentally Relevant Microbes to Inform Trace Metal Fate in the Environment

Advisor: Dr. Heileen Hsu-Kim

Organic thiol groups are important complexing ligands for chalcophile metals, which include contaminants like mercury and cadmium, as well as commodity metals like gold and silver. The highly stable complexes that thiols form with chalcophile metals have important implications for the transport, bioavailability, speciation, and distribution of these metals in the environment. However, there have been difficulties associated with measuring concentrations of thiol sites on complex biomaterials. Several methods have been developed to probe these sites, including fluorescence spectroscopy based methods and potentiometric titration based methods. In this research, fluorescence spectroscopy and potentiometric titration based methods are compared in their efficacy of estimating thiol concentrations on the cell envelope of Gram-positive bacteria, B. subtilis. Furthermore, several methods for modeling the surface protonation of B. subtilis to estimate thiol concentrations are compared, including Protofit, an open source protonation optimization software, and a Python code written to improve the Protofit algorithm’s capabilities. It was found that the inner-filter effect for the fluorescence spectroscopy method substantially inhibited accurate thiol measurements from being made. Additionally, surface complexation modeling choices in the potentiometric titration method significantly influence the optimized thiol concentration, especially the choice of modeling bacterial cell envelopes with three or four discrete acidic sites.

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