Celebration of Scholarship and Creative Activity 2023
Manganese Homeostasis in Bacteria: Identification of the Protein Bonding between the Small Protein MntS and Manganese Exporter MntP in E. coli
Manganese (Mn) is an essential trace nutrient for organisms because of its role in cofactoring enzymes and providing protection against reactive oxygen species (ROS). Many bacteria require manganese to form pathogenic or symbiotic interactions with eukaryotic host cells, in which maintaining Mn homeostasis is critical for bacterial survival. Virulent strains of bacteria sequester metal ions from their host species; however, excess intake of these ions lead to toxicity to the bacteria. To better understand regulation of intracellular metal concentrations, our research group focused on the interactions of proteins responsible for Mn homeostasis in Escherichia coli (E. coli), specifically focusing on small protein interactions thought to have important regulatory roles. We investigated the small protein MntS, only 42 amino acids long, with its interaction between MntP, a Mn exporter protein. To determine how these two proteins interact, we used an unbiased approach where PCR mutagenesis generated a library of random mutations in MntS. We then used a genetic screen to identify MntS mutants that were defective in binding to MntP. The identified mutants from the random screen were transformed into E.coli cells using site-directed PCR mutagenesis and were further tested to quantify the defects of protein-protein interactions through two-hybrid and Mn sensitivity assays. Indeed, using these two assays, we have demonstrated decreased interactions between MntS and MntP in multiple specific amino acid mutations. These results provide deeper insight into E. coli’s metal ion regulation, which in the long-run could allow better treatment of bacterial infections by targeting metal ion regulators.
Lindsay's partner Magdalyn Introduces The Project
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