Investigating the Role of ppsA in ATP Production and Phage Replication in E. coli

Abstract

Antibiotic resistance is one of the most pressing health crises of our time, threatening our ability to treat even common bacterial infections. In response to increasing antibiotic-resistant bacterial infections, phage therapy has become a promising alternative to traditional antibiotics. Bacteriophages infect bacterial cells in order to exploit the host’s metabolic products for their own growth and replication, ultimately killing the bacterial cell. That is, phages infect only bacterial cells, therefore treating infection without interfering with human cells.

Our project aimed to investigate cell growth and bacteriophage replication in E. coli when the ppsA gene is removed to provide further insight into phage therapy. The gene ppsA is located in the gluconeogenesis I pathway, and encodes the enzyme phosphoenolpyruvate synthetase (ppsA). PpsA catalyzes the reaction that produces phosphoenolpyruvate (PEP), a precursor for glycolysis involved in the production of ATP. Comparative growth assays, lysis curves, comparative plaque assays, and an ATP quantification assay were performed to quantify the growth, viral replication, and ATP levels of parent and ΔppsA strains.

ΔppsA exhibited a slower growth rate than wild type in LB and M9 minimal media, replicating previous findings that deleting ppsA hinders E. coli growth. ΔppsA also experienced reduced T2 and T4r bacteriophage replication, a novel finding. Results of an ATP quantification assay were produced but inconclusive as to if ATP availability was directly impacted by ΔppsA. Overall, a direct link between ppsA and effective bacteriophage replication was confirmed. This link between ppsA and bacteriophage growth indicates that future phage therapy strategies may benefit from targeting bacterial metabolic pathways to improve treatments. Future work will be needed to determine the mechanisms behind this relationship, especially the role of ATP availability in ΔppsA.