Question

Considering the Metabolic pathways of Pseudomonas aeruginosa:

A. Briefly evaluate the metabolism of the organism. How do they make PMF, ATP and reducing power? Do they have a broad or a narrow metabolic capacity?

B. How would deleting the TCA cycle affect this organism?

C. What about a mutation in the quinol binding site of the bc1 complex (Complex III) so that it could not accept a quinol?

D. What about a mutation in the gene that encodes glyceraldehyde-3-P dehydrogenase (the enzyme that catalyzes the oxidation of G-3-P)?

Answer 1

Protein F--Since P. aeruginosa is a Gram-negative microbe, it has an outer membrane which contains Protein F (OprF). OprF functions as a porin, allowing certain molecules and ions to come into the cells, and as a structural protein, maintaining the bacterial cell shape. Because OprF provides P. aeruginosa outer membrane with an exclusion limit of 500 Da, it lowers the permeability of the outer membrane, a property that is desired because it would decrease the intake of harmful substances into the cell and give P. aeruginosa a high resistance to antibiotics .

Flagellum and Pili--P. aeruginosa uses its single and polar flagellum to move around and to display chemotaxis to useful molecules, like sugars. Its strains either have a-type or b-type of flagella, a classification that is based primarily on the size and antigenicity of the flagellin subunit. The flagellum is very important during the early stages of infection, for it can attach to and invade tissues of the hosts . Similarly to its flagellum, P. aeruginosa pili contribute greatly to its ability to adhere to mucosal surfaces and epithelial cells. Specifically, it is the pili’s tip that is responsible for the adherence to the host cell surface. P. aeruginosa have N-methyl-phenyl-alanine (NMePhe) or type IV pili . The pili are characterized as long polar filaments made up of homopolymers from the protein pilin, which is encoded by the pilA gene. Overall, P. aeruginosa flagellum and pili have similar functionality (for attachment) and structure (both are filamentous structures on the surface of the cell), and their motility is controlled by RpoN, especially during initial attachment to the human host and under low nutrient conditions.

Pseudomonas aeruginosa Scanning Electron Micrograph. From the Centers for Disease Control and Prevention(CDC)

When infecting its host, P. aeruginosa is starved for iron because iron deprivation of an infecting pathogen is the key part in the humans’ innate defense mechanism. To overcome this challenge, P. aeruginosa synthesizes two siderophores: pyochelin and pyoverdin. P. aeruginosa then secrets these sideophores to the exterior of the cell, where they bind tightly to iron and bring the iron back into the cell. Additionally, P. aeruginosa can also use iron from enterobactin, a special siderophore produced by E. coli for iron transport, to satisfy its iron need.

P. aeruginosa is a facultative aerobe; its preferred metabolism is respiration. It gains energy by transferring electrons from glucose, a reduced substrate, to oxygen, the final electron acceptor. The breakdown of glucose requires it to oxidize to gluconate in the periplasm, then it will be brought inside the inner membrane by a specific energy-dependent gluconate uptake system. Once inside, gluconate is phosphorylated to 6-P-gluconate, which will enter the central metabolism to produce energy for the cell . When P. aeruginosa is in anaerobic conditions, however, P. aeruginosa uses nitrate as a terminal electron acceptor. Under oxidative-stress conditions, P. aeruginosa synthesizes Fe- or Mn- containing superoxide dismutase (SOD) enzymes, which catalyze the very reactive O- to H2O2 and O2. It also detoxifies H2O2 to O2 and H2O by using catalase.