Biochemical and physiological characterization of the three NADH dehydrogenases in Pseudomonas aeruginosa

Abstract

Pseudomonas aeruginosa is a Gram-negative opportunistic pathogen that is present in a wide variety of environments. Immune compromised patients readily acquire P. aeruginosa. After infection, this bacterium typically outcompetes all other bacteria in their environment and persists, creating a chronic infection that is extremely difficult to eradicate. P. aeruginosa infections are prevalent because of their incredible ability to adapt. Much of this adaptability stems from the flexibility of the respiratory pathway of this bacterium.

The aerobic electron transport chain in P. aeruginosa is extremely complex, consisting of a branched network of almost all known classes of respiratory enzymes, including three NADH dehydrogenases: the Na+-translocating NADH:ubiquinone oxidoreductase (Na+-NQR), the H+-translocating NADH:ubiquinone reductase (Complex I/NUO), and a non-ion pumping NADH dehydrogenase (NDH2). Na+-NQR and NUO pump Na+ and H+, respectively, and contribute to the electrochemical gradient used by the cell for metabolic work. Many bacterial species have a single type of ion-pumping NADH dehydrogenase, either Na+-NQR or NUO. P. aeruginosa, on the other hand, has both enzymes, which allows the cell to generate both H+ and a Na+ gradients directly.

This project examines the role of the NADH dehydrogenases in respiration and virulence in P. aeruginosa. We have found that all three of the NADH dehydrogenases (Na+-NQR, NUO and NDH2) are active during both exponential and stationary phases of growth, but are regulated differently, and do not compensate for each other when one is eliminated. Inactivating any of the NADH dehydrogenases produces different, but profound effects on virulence, pyocyanin production, biofilm formation, antibiotic resistance and NADH availability in the cell. This leads to the conclusion that regulation of the NADH dehydrogenases helps to control the onset of virulence and stationary phase-like behavior in P. aeruginosa.