Pathogenic bacteria require iron for their survival and ability to cause infection. Many bacterial pathogens have evolved sophisticated systems to utilize heme as a primary source of iron. The opportunistic pathogen P. aeruginosa is an increasingly common cause of nosocomial infections with high morbidity associated with such infections. In addition to their natural resistance to many antibiotics the alarming increase in such infections highlights the need for alternate antimicrobial strategies. In previous biochemical and biophysical studies we have characterized the intracellular heme trafficking protein PhuS and the iron-regulated heme oxygenase HemO and have further shown they are critical for heme utilization and virulence.
The specific aims of the proposal are to further elucidate the mechanism of extracellular heme utilization and regulation as it relates to virulence. This will be achieved through; i) Elucidating the intracellular heme trafficking pathway and defining the role of biliverdin IX-delta and -beta isomers in the extra-cellular feedback regulation of heme uptake through genetic, biochemical, and metabolomics approaches. ii) Defining the mechanism of heme transfer from PhuS to HemO through a combination of site-directed mutagenesis, biochemical and biophysical studies and iii) Determining the role of the regulatory small RNA PrrH in heme uptake and utilization. These studies will provide new insights into heme and iron homeostasis in bacterial pathogens. We will through this proposal identify new heme metabolic and regulatory pathways as novel antimicrobial targets and advance the current understanding of heme transfer mechanisms.

Public Health Relevance

The rapid rise in antibiotic resistance in the past two decades increases the need for alternative therapeutic strategies. A current approach to developing new antibiotics is to target non-essential functions that reduce virulence but decrease the selective pressure on the organism to undergo mutation. Heme utilization provides such a target and elucidation of the molecular mechanisms of heme trafficking and utilization will aid in identifying novel therapeutic targets.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
3R01AI102883-04S1
Application #
9223037
Study Section
Special Emphasis Panel (ZRG1-BCMB-B (02))
Program Officer
Ernst, Nancy Lewis
Project Start
2012-12-01
Project End
2017-11-30
Budget Start
2016-02-12
Budget End
2016-11-30
Support Year
4
Fiscal Year
2016
Total Cost
$38,887
Indirect Cost
$13,636
Name
University of Maryland Baltimore
Department
Pharmacology
Type
Schools of Pharmacy
DUNS #
188435911
City
Baltimore
State
MD
Country
United States
Zip Code
21201
Huang, Weiliang; Wilks, Angela (2017) A rapid seamless method for gene knockout in Pseudomonas aeruginosa. BMC Microbiol 17:199
MouriƱo, Susana; Giardina, Bennett J; Reyes-Caballero, Hermes et al. (2016) Metabolite-driven Regulation of Heme Uptake by the Biliverdin IX?/?-Selective Heme Oxygenase (HemO) of Pseudomonas aeruginosa. J Biol Chem 291:20503-15
Heinzl, Geoffrey A; Huang, Weiliang; Yu, Wenbo et al. (2016) Iminoguanidines as Allosteric Inhibitors of the Iron-Regulated Heme Oxygenase (HemO) of Pseudomonas aeruginosa. J Med Chem 59:6929-42
Smith, Aaron D; Modi, Anuja R; Sun, Shengfang et al. (2015) Spectroscopic Determination of Distinct Heme Ligands in Outer-Membrane Receptors PhuR and HasR of Pseudomonas aeruginosa. Biochemistry 54:2601-12
Smith, Aaron D; Wilks, Angela (2015) Differential contributions of the outer membrane receptors PhuR and HasR to heme acquisition in Pseudomonas aeruginosa. J Biol Chem 290:7756-66
Reinhart, Alexandria A; Powell, Daniel A; Nguyen, Angela T et al. (2015) The prrF-encoded small regulatory RNAs are required for iron homeostasis and virulence of Pseudomonas aeruginosa. Infect Immun 83:863-75
Nguyen, Angela T; O'Neill, Maura J; Watts, Annabelle M et al. (2014) Adaptation of iron homeostasis pathways by a Pseudomonas aeruginosa pyoverdine mutant in the cystic fibrosis lung. J Bacteriol 196:2265-76
Wilks, Angela; Ikeda-Saito, Masao (2014) Heme utilization by pathogenic bacteria: not all pathways lead to biliverdin. Acc Chem Res 47:2291-8
Wilks, Angela; Heinzl, Geoffrey (2014) Heme oxygenation and the widening paradigm of heme degradation. Arch Biochem Biophys 544:87-95
O'Neill, Maura J; Wilks, Angela (2013) The P. aeruginosa heme binding protein PhuS is a heme oxygenase titratable regulator of heme uptake. ACS Chem Biol 8:1794-802

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