Protein Interactions in the Utilization of Iron by Bacteria

University Of Kansas Center For Research Inc, Lawrence KS

Investigators

Abstract

Intellectual Merit: Iron is a required nutrient involved in fundamental cellular processes, from oxygen storage and transport to respiration and DNA repair. Pathogenic bacteria are equipped with iron uptake mechanisms that enable them to utilize host-iron sequestered by lactoferrin, transferrin or bound to heme, and therefore overcome the low-iron concentrations encountered in their host organisms. Much less is known about the fate of iron once it reaches the bacterial cytosol. Ferritin molecules function as dynamic regulators of cytosolic iron concentrations, but despite their importance in iron homeostasis, little is known about the processes that deliver Fe2+ for storage or the signals that prompt its release for safe integration in metabolism. There are two ferritin-like molecules in P. aeruginosa: a bacterioferritin (BfrB), which assembles from 24 identical subunits and binds 12 heme molecules, and a bacterial ferritin (FtnA), which is also assembled from 24 identical subunits but does not bind heme. It has recently been discovered that release of iron from BfrB requires electrons to be mediated by a ferredoxin (Bfd) and a ferredoxin reductase (FPR), whereas release of iron from FtnA requires that electrons be mediated only by FPR. These findings provide a unique platform to study how protein structure and protein-protein interactions contribute to the regulation of cytosolic iron in pathogenic bacteria. In order to investigate these protein-protein interactions a multidisciplinary approach will be undertaken which includes biochemistry, NMR spectroscopy, X-ray crystallography and computational biology. The specific goals are: 1) Investigate and structurally determine the protein-protein interactions that regulate the outflow of iron from the interior cavities of BfrB and FtnA. (2) Challenge a hypothesis that Fe2+ exits the BfrB and FtnA interior cavities via the four-fold pores formed in their structures at the intersection of 4-subunits. Broader Impacts: The multidisciplinary nature of the project will provide many opportunities for students at all academic levels. The integration of efforts in the Rivera, Vakser and Lovell laboratories (all located at the University of Kansas) via monthly joint group meetings will expose the students to the intellectually diverse atmosphere that is necessary to nurture multidisciplinary research. This experience, coupled to the ethnically rich environment present in the three labs (Asian, Caucasian, Hispanic; ~40/60 female/male) will prepare the graduate students well for the multifaceted work environment they are likely to encounter after graduation. Just as important, the collaborative spirit of this project will illustrate to students the benefits of a broad-based approach for (1) solving complex problems; (2) more effectively putting the results obtained in one laboratory into its proper context; and thus (3) improving the prospect that the research will impact the greater scientific community, and ultimately benefit the general public. This project is jointly supported by the Biomolecular Dynamics, Structure and Function Cluster in the Division of Molecular and Cellular Biosciences and the Chemistry of Life Processes program in the Chemistry Division.