Iron is an essential nutrient for most organisms, including pathogenic bacteria. Pathogenic bacteria attempting to colonize humans are confronted with extremely low concentrations of free iron. Consequently, many pathogens have evolved sophisticated mechanisms for iron acquisition, including the utilization of heme-iron. Moreover, it has been recently shown that during the early stages of infection Staphylococcus aureus prefers iron from heme. Thus, it is possible that targeting paths used by pathogenic bacteria to assimilate iron and heme-iron from their host is a viable approach to de development of new antibiotics. Many of the proteins involved in heme uptake and heme utilization in the opportunistic pathogen Pseudomonas aeruginosa have designated functions. However, the structure, dynamics and inter-protein interactions that facilitate host-heme capture, internalization and degradation in the cytosol are largely unknown. In this application we propose to contribute to fill this gap by studying the structure, function, dynamics and association of the soluble proteins that aid in the capture of heme from hemoglobin and help degrade it in the cytosol of P. aeruginosa. Important outcomes of the proposed studies are: (1) Biochemical and structural characterization of two previously unknown electron transport proteins (Bfd and Fpr), which we hypothesize function to deliver the 7 electrons needed by heme oxygenase to cleave the heme and release its iron in the cytosol of P. aeruginosa. We also plan to investigate the protein-protein interactions that facilitate electron transfer from Bfd to heme oxygenase to support the degradation of heme. (2) Structural characterization of HasAp, a secreted heme binding protein capable of capturing heme from hemoglobin and delivering it to the outer membrane receptor for internalization. The acquired structural information of HasAp will be used to define its interactions with hemoglobin (3) Characterization of how polypeptide dynamics contribute to the heme oxidation activity of heme oxygenase from P. aeruginosa, which at this point is better understood largely due to work supported in the expiring funding cycle. The information learned from these investigations is expected to provide several entry points for the future design of novel therapeutic strategies aimed at interfering with heme uptake and degradation in P. aeruginosa. ? ? ?

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
2R01GM050503-10A2
Application #
7199450
Study Section
Special Emphasis Panel (ZRG1-BCMB-B (02))
Program Officer
Preusch, Peter C
Project Start
1995-04-01
Project End
2010-08-31
Budget Start
2006-09-15
Budget End
2007-08-31
Support Year
10
Fiscal Year
2006
Total Cost
$259,950
Indirect Cost
Name
University of Kansas Lawrence
Department
Chemistry
Type
Schools of Arts and Sciences
DUNS #
076248616
City
Lawrence
State
KS
Country
United States
Zip Code
66045
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