An integrated network analysis reveals that nitric oxide reductase prevents metabolic cycling of nitric oxide by Pseudomonas aeruginosa |
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| Institution: | 1. Department of Chemical and Biological Engineering, Princeton University, Princeton, NJ 08544, USA;2. Department of Molecular Biology, Princeton University, Princeton, NJ 08544, USA;3. Rutgers Robert Wood Johnson Medical School, Piscataway, NJ 08854, USA;1. Department of Chemical Engineering, Villanova University, Villanova, PA;2. The Henry M. Jackson Foundation for the Advancement of Military Medicine, Bethesda, MD;3. Departments of Chemical Engineering, Pennsylvania State University, University Park, PA;4. The Center for Nonlinear Dynamics & Control (CENDAC), Villanova University, Villanova, PA;5. Center for the Advancement of Sustainability in Engineering (VCASE), Villanova University, Villanova, PA;1. Joint BioEnergy Institute, 5885 Hollis Street, Emeryville, CA 94608, USA;2. Biological Systems & Engineering Division, Lawrence Berkeley National Laboratory, Berkeley, CA 94720, USA;3. Department of Bioengineering, University of California, Berkeley, CA 94720, USA;4. Department of Chemical and Biomolecular Engineering, University of California, Berkeley, CA 94720, USA;5. The Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark, Denmark;1. School of Mechanical Engineering, University of Science and Technology Beijing, Beijing 100083, China;2. School of Engineering and Applied Sciences, Harvard University, 02138 Cambridge, MA, USA;1. College of Environment, Zhejiang University of Technology, Hangzhou 310032, China;2. Key Laboratory of Pollution Exposure and Health Intervention of Zhejiang Province, Interdisciplinary Research Academy, Zhejiang Shuren University, Hangzhou 310015, China;3. Zhejiang Envrionmental Monitoring Engineering Co., Ltd, China;4. Research Institute of Physical and Chemical Problems, Belarusian State University, Minsk 220030, Belarus |
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| Abstract: | Nitric oxide (NO) is a chemical weapon within the arsenal of immune cells, but is also generated endogenously by different bacteria. Pseudomonas aeruginosa are pathogens that contain an NO-generating nitrite (NO2?) reductase (NirS), and NO has been shown to influence their virulence. Interestingly, P. aeruginosa also contain NO dioxygenase (Fhp) and nitrate (NO3?) reductases, which together with NirS provide the potential for NO to be metabolically cycled (NO→NO3?→NO2?→NO). Deeper understanding of NO metabolism in P. aeruginosa will increase knowledge of its pathogenesis, and computational models have proven to be useful tools for the quantitative dissection of NO biochemical networks. Here we developed such a model for P. aeruginosa and confirmed its predictive accuracy with measurements of NO, O2, NO2?, and NO3? in mutant cultures devoid of Fhp or NorCB (NO reductase) activity. Using the model, we assessed whether NO was metabolically cycled in aerobic P. aeruginosa cultures. Calculated fluxes indicated a bottleneck at NO3?, which was relieved upon O2 depletion. As cell growth depleted dissolved O2 levels, NO3? was converted to NO2? at near-stoichiometric levels, whereas NO2? consumption did not coincide with NO or NO3? accumulation. Assimilatory NO2? reductase (NirBD) or NorCB activity could have prevented NO cycling, and experiments with ΔnirB, ΔnirS, and ΔnorC showed that NorCB was responsible for loss of flux from the cycle. Collectively, this work provides a computational tool to analyze NO metabolism in P. aeruginosa, and establishes that P. aeruginosa use NorCB to prevent metabolic cycling of NO. |
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| Keywords: | Metabolic cycle Kinetic model Oscillations NO reductase Fhp Denitrification |
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