The comparative role of molecular and nitrate-oxygen in the dissimilation of glucose Public Deposited

http://ir.library.oregonstate.edu/concern/graduate_thesis_or_dissertations/vx021h229

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  • When nitrate is used in a biological electron transport system as the final acceptor of hydrogen to form water, the system is characterized as "Nitrate Respiration." However, the quantitative expression of the over-all participation of NO₃⁻(0) in the oxidation of glucose by denitrifiers such as Pseudomonas stutzeri deserves clarification. In addition, the identity of the pathways of glucose breakdown that operate with NO₃⁻ are not understood nor is the dual role of molecular and nitrate oxygen. It is the purpose of the present study to provide clarification of the role of nitrate in the utilization of glucose by P. stutzeri. A strain of P. stutzeri was used throughout the entire study. The culture was maintained on nitrate broth or agar. However, a semi-synthetic basal medium was used for the experimental studies. The individual experiments were carried out using both an electrolytic respirometer and a microscale radiorespirometric apparatus. Each assembly provided a closed system thereby allowing for the determination of any evolved gases. Gases were detected using a Beckman G C-2 chromatograph. Analysis for nitrites, nitrates and the ammonium ions were determined by established chemical procedures. Radioactivity of evolved C¹⁴O₂ was determined by counting of barium carbonate planchets and also by using the Tri-carb Packard Scintillation Spectrometer. Appropriate calculations for specific activity, specific yield and relative specific activity were used. The results of the present study clarify several points relating to the role of nitrate and molecular oxygen as terminal hydrogen acceptors in P. stutzeri. First the use of high nitrate levels (500-1000 ppm NO₃⁻) markedly inhibits the uptake of molecular oxygen and the evolution of carbon dioxide. The nitrate inhibition effect progressively increases with increasing concentrations of nitrate. However, the observed inhibition decrease with time is at 1000 ppm NO₃⁻ no inhibition is observed after 48 hours incubation. At relatively low concentrations of nitrate (250-500 ppm NO₃⁻) both NO₃⁻(O) and molecular (O) act as dual electron acceptors. In the latter instances, nitrate contributes one-third and molecular oxygen two-thirds of the total oxygen required for the production of one millimole of carbon dioxide. A final observation relates to the pathway of glucose catabolism. High nitrate levels effect a decrease in the normal phosphogluconate or direct oxidative route characteristic of pure aerobic respiration in P. stutzeri. The shift in the glucose breakdown pathway appears to go over to classical glycolysis. Thus, "nitrate respiration" in P. stutzeri appears to make use of the EMP pathway rather than the route to pentose formation. This aspect of the study is presently being given additional emphasis.
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