Siderophores are low molecular weight iron chelators produced by bacteria to scavenge iron from the environment, and are frequently required for bacterial virulence. The long-term goal of the research in this laboratory is to provide a detailed mechanistic and structural understanding of enzymes in the siderophore biosynthetic pathways that are targets for drug discovery. Toward that end, we propose to investigate the nonribosomal peptide synthetase accessory enzymes that are involved in the production of the siderophores pyochelin and pyoverdin from the bacteria P. aeruginosa. Two of the four enzymes to be studied, the isochorismate synthase (PchA) and isochorismate-pyruvate lyase (PchB), are ideal model systems for making fundamental advances in understanding alternate reaction pathways for catalysis. The studies of the ornithine hydroxylase (PvdA) show promise for novel flavin biochemistry, and the first characterization of the hydroxyornithine transformylase (PvdF) is proposed. We will use a multifaceted approach that includes enzyme kinetic analysis, structural biology and computational enzymology. Narrative Pseudomonas aeruginosa is a dangerous pathogen that is a common cause of infections in susceptible hosts, including cancer and AIDS patients, those with immune deficiencies, cystic fibrosis patients, burn patients and other at risk individuals including infants. The goal of this work is to provide a fundamental understanding of four enzymes that promote virulence in P. aeruginosa so that this information can be exploited to generate new antimicrobial drugs.

Public Health Relevance

Pseudomonas aeruginosa is a dangerous pathogen that is a common cause of infections in susceptible hosts, including cancer and AIDS patients, those with immune deficiencies, cystic fibrosis patients, burn patients and other at risk individuals including infants. The goal of this work is to provide a fundamental understanding of four enzymes that promote virulence in P. aeruginosa so that this information can be exploited to generate new antimicrobial drugs.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Scientist Development Award - Research (K02)
Project #
5K02AI093675-03
Application #
8431451
Study Section
Microbiology and Infectious Diseases B Subcommittee (MID)
Program Officer
Taylor, Christopher E,
Project Start
2011-03-01
Project End
2015-02-28
Budget Start
2013-03-01
Budget End
2014-02-28
Support Year
3
Fiscal Year
2013
Total Cost
$106,920
Indirect Cost
$7,920
Name
University of Kansas Lawrence
Department
Biology
Type
Schools of Arts and Sciences
DUNS #
076248616
City
Lawrence
State
KS
Country
United States
Zip Code
66045
Meneely, Kathleen M; Luo, Qianyi; Riley, Andrew P et al. (2014) Expanding the results of a high throughput screen against an isochorismate-pyruvate lyase to enzymes of a similar scaffold or mechanism. Bioorg Med Chem 22:5961-9
Chilton, Annemarie S; Ellis, Ashley L; Lamb, Audrey L (2014) Structure of an Aspergillus fumigatus old yellow enzyme (EasA) involved in ergot alkaloid biosynthesis. Acta Crystallogr F Struct Biol Commun 70:1328-32
Frederick, Rosanne E; Ojha, Sunil; Lamb, Audrey et al. (2014) How pH modulates the reactivity and selectivity of a siderophore-associated flavin monooxygenase. Biochemistry 53:2007-16
Meneely, Kathleen M; Luo, Qianyi; Lamb, Audrey L (2013) Redesign of MST enzymes to target lyase activity instead promotes mutase and dehydratase activities. Arch Biochem Biophys 539:70-80
Meneely, Kathleen M; Luo, Qianyi; Dhar, Prajnaparamita et al. (2013) Lysine221 is the general base residue of the isochorismate synthase from Pseudomonas aeruginosa (PchA) in a reaction that is diffusion limited. Arch Biochem Biophys 538:49-56
Olucha, Jose; Lamb, Audrey L (2011) Mechanistic and structural studies of the N-hydroxylating flavoprotein monooxygenases. Bioorg Chem 39:171-7
Lamb, Audrey L (2011) Pericyclic reactions catalyzed by chorismate-utilizing enzymes. Biochemistry 50:7476-83
Olucha, Jose; Ouellette, Andrew N; Luo, Qianyi et al. (2011) pH Dependence of catalysis by Pseudomonas aeruginosa isochorismate-pyruvate lyase: implications for transition state stabilization and the role of lysine 42. Biochemistry 50:7198-207
Olucha, Jose; Meneely, Kathleen M; Chilton, Annemarie S et al. (2011) Two structures of an N-hydroxylating flavoprotein monooxygenase: ornithine hydroxylase from Pseudomonas aeruginosa. J Biol Chem 286:31789-98