Pyruvate Carboxylase Plays a Crucial Role in Carbon Metabolism of Extra- and Intracellularly ReplicatingListeria monocytogenes

Abstract

ABSTRACTThe human pathogenL. monocytogenesis a facultatively intracellular bacterium that survives and replicates in the cytosol of many mammalian cells. The listerial metabolism, especially under intracellular conditions, is still poorly understood. Recent studies analyzed the carbon metabolism ofL. monocytogenesby the13C isotopologue perturbation method in a defined minimal medium containing [U-13C6]glucose. It was shown that these bacteria produce oxaloacetate mainly by carboxylation of pyruvate due to an incomplete tricarboxylic acid cycle. Here, we report that apycAinsertion mutant defective in pyruvate carboxylase (PYC) still grows, albeit at a reduced rate, in brain heart infusion (BHI) medium but is unable to multiply in a defined minimal medium with glucose or glycerol as a carbon source. Aspartate and glutamate of thepycAmutant, in contrast to the wild-type strain, remain unlabeled when [U-13C6]glucose is added to BHI, indicating that the PYC-catalyzed carboxylation of pyruvate is the predominant reaction leading to oxaloacetate inL. monocytogenes. ThepycAmutant is also unable to replicate in mammalian cells and exhibits high virulence attenuation in the mouse sepsis model.