Lactobacillus sakei is a lactic acid bacterium belonging to the natural flora of fresh and vacuum-packed meat, and is used as the starter for manufacturing fermented sausages. This species is now being studied at the genetic level. We investigated uracil prototrophy of strain 23K in order to validate the proteomic approach to study metabolism regulations. Cells grown without uracil had lower growth rates than with uracil. Protein analysis by 2D gel electrophoresis showed that at least three polypeptides were specifically induced in the absence of uracil. Two of these polypeptides were identified as orotate phosphoribosyl transferase, catalyzing the fifth step of pyrimidine biosynthesis, and PyrR, the transcriptional regulator of the pyr operon, respectively.