The metabolism of 2-furoic acid by Pseudomonas F2

1. Pseudomonas F2 isolated by enrichment culture on 2-furoic acid and grown with it as carbon source oxidized the compound with a Q_o2 of 170μl./mg. dry wt./hr. and the overall consumption of 2·5μmoles of oxygen/μmole of substrate. 2. In the presence of 1mm-sodium arsenite, oxygen uptake was restricted to 0·54μmole/μmole of 2-furoate oxidized, with the formation of 0·86μmole of 2-oxoglutarate/μmole of 2-furoate. 3. Cell suspensions, disrupted in a French pressure cell and centrifuged at 27000g, yielded supernatants capable of catalysing the slow oxidation of 2-furoate (0·17μmole/mg. of protein/hr.). 4. Fractionation of 27000g supernatants at 200000g yielded a soluble enzyme fraction capable of catalysing the oxidation of 2-furoate only in the presence of added 200000g pellet or of Methylene Blue. 5. The 2-furoate-stimulated uptake of oxygen or the anaerobic reduction of Methylene Blue by dialysed 27000g supernatant required the addition of ATP and CoA, and the rate of oxygen uptake was further enhanced by the addition of magnesium chloride and NAD⁺. 6. The role of ATP and CoA in the formation of 2-furoyl-CoA was demonstrated by the accumulation of 2-furoylhydroxamic acid in the presence of hydroxylamine. 7. Dialysed 200000g supernatant, treated with Dowex 1, required the addition of ATP, CoA and Methylene Blue before it could oxidize 2-furoate to 2-oxoglutarate, which was trapped in unitary stoicheiometric yield as its phenylhydrazone. Magnesium chloride and NAD⁺ were not stimulatory in this system. The oxidation of 2-furoate to 2-oxoglutarate was not inhibited by substrate analogues, metal ion-chelating agents, thiol-active compounds or inhibitors of cytochrome-mediated electron transport. 8. No evidence was obtained for the intervention of 2,5-dioxovalerate as an intermediate in 2-oxoglutarate formation.