A safe food system depends on maintenance of a safe water system. Both chemical (e.g. microplastics) and biological (e.g. microbial pathogens) hazards present in water can be harmful to agricultural practices and direct consumption. My dissertation research employs principles of protein engineering and synthetic biology in practical solutions to challenges in water safety. I engineered bacteriophage capsid chemistry to enhance rapid bacterial detection in drinking water and manipulated enzyme chemistry to degrade microplastics in agricultural water. To meet regulatory standards, drinking water must have 0 detectable E. coli (a standard indicator of fecal contamination) per sample. Detection systems must therefore reliably indicate 1 colony-forming unit (cfu) of E. coli per standard 100 ml sample, a size difference of 1015 between sample and analyte. Bacteriophages, which specifically bind a tunable range of bacteria, are an ideal biorecognition element in ultrasensitive bacterial detection platforms. I built a T4 bacteriophage-based biosensor using site-specific artificial amino acid incorporation to enable bioorthogonal azide-alkyne “click chemistry” conjugation to magnetic nanoparticles. By combining magnetic separation with the native phage bacterial attachment system, this sensor concentrates the E. coli within the water sample 1000-fold. This signal amplification enables single cfu detection within 7 hours, significantly faster than the 1-2 days required for the standard method. Microplastics (MPs) are plastic particles and fibers &lt;5 mm in their longest dimension. As plastic has become ubiquitous in modern life, MPs have accumulated in a range of environments. MPs can enter agricultural spaces, via treated wastewater (effluent). While the treatment process should render effluent, a common irrigation source, hazard-free, current treatment practices do not adequately address MP contamination. I modified the amino acid composition of a natural plastic-degrading enzyme, PETase, to enable in situ MP biodegradation in wastewater treatment plants. This enzyme degrades a common plastic into soluble monomers readily metabolized by bacteria. Using a combination of computational design, site-directed mutagenesis, and combinatorial engineering, I increased enzymatic activity by over 300%. Overall, my work shows that precise manipulation of amino acid chemistry can transform naturally occurring biologics into potent, practical tools for enhancing water safety.

Tailoring protein chemistry for in situ water safety applications: rapid pathogen detection and microplastic degradation

ACS Spring 2022

The official archive and repository for all ACS Meetings & Expositions abstracts and select posters or presentations.