No abstract

In a previous publication (1) evidence was presented impficating succinate and pyruvate quantitatively in the oxidative breakdown of acetic acid to CO2 and water by acetate-adapted strains of Escherichia coli, while a-ketoglutaric acid was eliminated as a possible intermediate. In addition, fumarate, malate, and oxalacetate have also been shown, qualitatively at least, to participate as intermediates in the oxidation of the C2 fatty acid. However, the occurrence of an unexpected side reaction, involving appreciable reduction of C4-dicarboxylic acid products of succinate oxidation back to succinate, complicated the interpretations of most experiments performed using carrier malate, fumarate, or oxalacetate.In this paper detailed data are presented which were obtained when 2-C 14-acetate was incubated in the presence of fumarate, malate, 0xalacetate, and a-ketoglutarate. Further, conditions will be described which reduce the unexpected reduction of the above mentioned C4-dicarboxylic acids back to succinate. From chemical analysis of the residual acids, CO~, and cell material, it was possible to show that when an appreciable fraction of acetate, as well as other substrates, was metabolized under high oxygen tension, there were incorporation and distribution of labeled carbon into fumarate and malate in the amount to be expected on the basis of a Thunberg-Knoop condensation cycle, whereas there was no comparable incorporation into a-ketoglutarate. These results plus those already published (1, 2) appear to exclude a tricarboxylic acid cycle as a major pathway for acetate oxidation in E. coll. Methods and MaterialsThe same strain of Esckerichia coli (E26) was used in these experiments as in the previous work. For all experiments cells were grown with constant aeration at 30°C. in a medium containing 1.5 per cent anhydrous sodium acetate, 0.4 per cent ammonium sulfate, 0.8 per cent KH2PO,, 0.07 per cent tryptone, and 20 per cent tap water (for 785

No abstract

Evidence against the occurrence of the Krebs oxidation cycle in bacterial respiration has been steadily accumulating. Escherichia coli and many other bacteria do not readily metabolize the three tricarboxylic acids. Aerobacter aerogenes will not readily attack citrate when measured manometrically unless the organism is grown in the presence of this acid as the sole source of carbon. Recently Lenti (1946) was able to show inhibition of succinic acid oxidation in E. coli by malonate, but the oxidation of pyruvate was not affected. Karlsson and Barker (1948) obtained evidence against the tricarboxylic acid cycle in Azotobacter agilis. There is, therefore, little support for the assumption that the cycle occurs in those bacteria whose intermediary metabolism has been studied in detail. On the other hand, many bacteria, including E. coli and A. aerogenes, oxidize succinate, fumarate, and malate, and reduce anaerobically oxalacetate to succinate, and this suggests that the Szent-Gy6rgyi system, which constitutes an integral part of the Krebs cycle, is operative in microorganisms. By the use of arsenious oxide and cyclohexanol it has been possible to show that glucose or pyruvate is oxidized aerobically as far as acetic acid without the mediation of the C4 dicarboxylic acids as catalytic hydrogen carriers. In other words, the possibility of pyruvic acid initially condensing with oxalacetate to form procitric acid or some other C7 intermediate in its oxidation scheme is ruled out. By applying the principle of "simultaneous adaptation" (Stanier, 1947), it was observed that acetate that arises from the breakdown of either glucose or pyruvic acid is further oxidized to CO2 and water without the mediation of cis-aconitate or a-ketoglutarate. These results may in turn be used as evidence against the occurrence of the tricarboxylic acid cycle in the organisms under consideration. METHODS E. coli or A. aerogenes was grown for 16 to 18 hours, with constant aeration, at 30 C in a medium containing 0.8 per cent substrate, 0.4 per cent (NH4)2SO4, 0.8 per cent KH2PO4, 0.2 per cent yeast extract or 0.07 per cent tryptone, and 20 per cent tap water, at an initial pH of 6.8 to 7.0. The cells were then harvested, washed several times with cold distilled water, and stored in the icebox, or first lyophilized and stored in the dry state. The results vary somewhat, particularly with respect to acetate oxidation by A. aerogenes, depending upon the treatment of the cells. The Barcroft-Warburg respirometer technique was used. The total volume per flask varied from 2.3 to 2.8 ml. Endogenous values were subtracted from those

No abstract