CAREER: Mapping the functional landscape of bacteriophage-host interactions at molecular resolution

While many bacteria are inoffensive or even beneficial to humans, some are harmful pathogens. Finding a way to kill the pathogenic bacteria without harming the beneficial microbes could prove a useful tool. Phage research has emerged as an exciting new frontier in microbiology due to its role in shaping microbiomes and as a potential therapy against drug-resistant bacterial infections. Phages identify and interact with their specific bacterial target through a structure known as the receptor-binding protein. While characterization of thousands of natural phages have occurred, limited studies have explored the molecular mechanisms that govern phage-host interactions through the receptor-binding protein. This research will apply genome engineering, high-throughput DNA sequencing, and viral metagenomics approaches to elucidate how protein sequences encode function. The research will be the basis for numerous viral-focused educational outreach activities, including for middle school classrooms, to expose the public to relevant, real-word issues in viral and bacterial biology. Receptor binding proteins mediate the interaction of a phage with its bacterial receptor(s) and thus constitute key determinants of a phage’s host range and activity (virulence). Phages exhibit high functional plasticity through genetic alterations of these proteins to adapt to new host environments. Despite extensive research to address the mechanisms associated with this interaction, it remains unclear how receptor binding protein mutations influence phage activity and host range. This work will examine the molecular rules of receptor binding proteins of coliphages, viruses that infect coliform bacteria, and will seek to exploit those rules to engineer synthetic coliphages with specificity for foodborne pathogens. In addition, the project will study receptor binding proteins from another group of phages that have a more complex receptor binding architecture that permits the phage to both enzymatically degrade the capsule that protects certain bacteria and also to bind to host receptors. To understand this process mechanistically, mutational and metagenomic scanning will be used to map and identify the function of every sequence in the receptor binding protein of a phage with the dual capability. These mutational studies will inform protein design strategies guided by machine learning to engineer new phage-host interactions. The project has the potential to create tools that could be broadly adopted by the research community to study sequence-function relationships in a variety of phage-host systems. This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.