UBC Theses and Dissertations
Metabolic activities of Pseudomonas Aeruginosa when grown on a 2-ketogluconic acid medium Hill, Robert William
The current concept of the pathway of the intermediate carbohydrate metabolism of P. aeruginosa ATCC 9027 is that glucose is oxidized to carbon dioxide and water by way of glucose, gluconic, 2-ketogluconic and pyruvic acids. The pathway of terminal respiration is the conventional Kreb's tricarboxylic acid cycle. However, the mechanism by which 2-ketogluconic acid is oxidized to pyruvic acid has not been elucidated. The first concern in the present work was to establish the accumulation of 2-ketogluconic acid during glucose oxidation by P. aeruginosa ATCC 9027 and also to produce enough calcium 2-ketogluconate to serve as substrate in studies of its oxidation. The initial problem on the metabolism of 2-ketogluconic acid grown cells of P. aeruginosa was to compare the enzymic pattern of these cells with cells grown on a glucose medium. The data obtained under these conditions confirmed the previous conclusions that 2-ketogluconic acid is in the direct pathway of glucose oxidation. Additional evidence for the minor role of the Embden-Meyer-hoff pathway in this organism was obtained when it was shown that cells grown on a 2-ketogluconic acid medium contained only traces of the enzyme aldolase. The amount of this enzyme detected would be sufficient to permit the glycolytic pathway to function as a synthesizing mechanism for pentoses or perhaps heptuloses but would not be sufficient to account for any major amount of glucose. The major problem of establishing the presence of intermediates occurring between 2-ketogluconic and pyruvic acids was approached from two points of view: first, by attempting to produce a cell preparation which had a limited ability to oxidize 2-ketogluconic acid and which would accumulate intermediates in a manner similar to that of cell preparations which accumulate 2-ketogluconic acid during glucose oxidation, and second, by the blocking of the enzyme system of 2-ketogluconic acid grown cells by an inhibitor at a point intermediate between 2-ketogluconic acid and pyruvic acid. Neither approach yielded positive results. Further studies were made on the basis of an early observation that cells which had been heated showed an impaired gluconic acid and 2-ketogluconic acid oxidizing activity. It was found subsequently that live cells heated to 55°C for at least ten minutes would oxidize glucose but not gluconic acid or 2-ketogluconic acid and that further heating nearly completely destroyed the ability of the cells to assimulate glucose. Lyophilized preparations of such heat treated cells still retained the ability to oxidize glucose or gluconic acid to 2-ketogluconic acid. This can be interpreted as indicating that 2-ketogluconic acid is on the direct oxidative pathway of glucose oxidation.
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