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    The Role of Iron Storage Proteins in Pseudomonas aeruginosa Bacterial Iron Homeostasis

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    Issue Date
    2016-08-31
    Author
    Eshelman, Kate
    Publisher
    University of Kansas
    Format
    133 pages
    Type
    Dissertation
    Degree Level
    Ph.D.
    Discipline
    Chemistry
    Rights
    Copyright held by the author.
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    Abstract
    Pseudomonas aeruginosa is a gram-negative bacterium that causes infections in immune compromised patients. There have been an increasing number of multi-drug resistant P. aeruginosa infections which is leading to the need to develop new targets for antibiotics. A potential new target is to disrupt iron homeostasis by disrupting the function of the iron storage protein, bacterioferritin B (BfrB). The structure and function of BfrB has been passionately studied in our lab, which has led to new understanding of iron uptake and iron release from BfrB. Iron mobilization from BfrB requires binding from the bacterioferritin-associated ferredoxin (Bfd), a process that our lab has demonstrated in vitro using X-ray crystallography, and binding studies. These studies also allowed the lab to determine the key residues in both proteins that stabilize the BfrB:Bfd complex. In my work, we have taken the insights from the in vitro studies and applied them to investigate the consequences of blocking the BfrB:Bfd interaction in P. aeruginosa cells. We first show that iron is essential to bacterial growth by testing the effects of an iron sequestering polymer developed in collaboration with Prof. Cory Berkland’s lab at the University of Kansas. The iron-sequestering polymer is capable of delaying bacterial growth and increasing the sensitivity of wild type (wt) P. aeruginosa to the antibiotics ciprofloxacin and gentamicin. I then studied cell growth and iron handling in response to mutating the bfrB gene (ΔbfrB), the bfd gene (Δbfd), or introducing a double mutation (E81A/L68A) in the bfrB gene in the chromosome of P. aeruginosa. From our previous in vitro studies, we predicted that E81/L68A BfrB mutant (herein denoted bfrB*) would not bind BfrB in P. aeruginosa cells. We demonstrate through these studies that BfrB and the BfrB:Bfd interaction are essential for iron homeostasis in P. aeruginosa. The structural dynamics of BfrB have also been analyzed. We show that by mutating residues in the B-pores of the protein, we affect the function of the relatively distant ferroxidase center, which in turn inhibits iron oxidation and uptake. We show that concerted motions linking the pores and the catalytic center are essential for the function of BfrB. Lastly, our lab is engaged in developing compounds for blocking the BfrB:Bfd interaction. I have developed assays to show the effect of these compounds on cell growth and survival, and demonstrated that the compounds being developed in the lab boost the killing activity of existing antibiotics against P. aeruginosa cells.
    URI
    http://hdl.handle.net/1808/25359
    Collections
    • Chemistry Dissertations and Theses [336]
    • Dissertations [4473]

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    Contact KU ScholarWorks
    785-864-8983
    KU Libraries
    1425 Jayhawk Blvd
    Lawrence, KS 66045
    785-864-8983

    KU Libraries
    1425 Jayhawk Blvd
    Lawrence, KS 66045
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    Contact KU ScholarWorks
    785-864-8983
    KU Libraries
    1425 Jayhawk Blvd
    Lawrence, KS 66045
    785-864-8983

    KU Libraries
    1425 Jayhawk Blvd
    Lawrence, KS 66045
    Image Credits
     

     

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