Citrate synthase, the first and rate-limiting enzyme of the tricarboxylic acid branch of the Krebs cycle, was shown to be required for de novo synthesis of glutamate and glutamine in Listeria monocytogenes. The citrate synthase (citZ) gene was found to be part of a complex operon with the upstream genes lmo1569 and lmo1568. The downstream isocitrate dehydrogenase (citC) gene appears to be part of the same operon as well. Two promoters were shown to drive citZ expression, a distal promoter located upstream of lmo1569 and a proximal promoter located upstream of the lmo1568 gene. Transcription of citZ from both promoters was regulated by CcpC by interaction with a single site; assays of transcription in vivo and assays of CcpC binding in vitro revealed that CcpC interacts with and represses the proximal promoter that drives expression of the lmo1568, citZ, and citC genes and, by binding to the same site, prevents read-through transcription from the distal, lmo1569 promoter. Expression of the lmo1568 operon was not affected by the carbon source but was repressed during growth in complex medium by addition of glutamine.