Acetate oxidation through a modified citric acid cycle in Propionibacterium freudenreichii

Abstract

Propionibacterium freudenreichii strain DSM 20271 was grown in a mineral medium containing 0.1% (w/v) yeast extract. Acetate was oxidized by growing cells with potassium hexacyanofen-ate as electron acceptot, which was oxidized by a three-electrode poised-potential system at a redox potential of +510 mV. Growth with acetate under these conditions followed linear rather than exponential kinetics, whereas growth with other substrates such as lactate under the same conditions was exponential. Cell-free extracts of P. freudenreichii cells grown with acetate contained all enzymes of the classical citric acid cycle except 2-oxoglutarate-oxidizing activity. No activity of anaplerotic reactions such as isocitrate lyase or malate synthase was found. Instead, moderate activities of glutamate decarboxylase, 4-aminobutyrate:2-oxoglutarate aminotransferase, and succinate semialdehyde dehydrogenase were detected. In short-term radiolabeling experiments with U-~4C-acetate, 4-aminobutyrate was identified as a major early intermediate in acetate oxidation by these cells. These findings allow the construction of a modified citric acid cycle that compensates the lack of 2-oxoglutarate dehydrogenase by a subcycle through glutamate, 4-aminobutyrate, and succinate semialdehyde. Lack of anaplerotic reactions explains subexponential growth kinetics during growth with acetate.