Microcin E492-producing bacteria secrete both unmodified and post-translationally modified microcins. The modification consists of a C-glucosylated linear trimer of N-(2,3-dihydroxybenzoyl)-L-serine, a catecholate siderophore related to salmochelins and enterobactin. We showed here that repression of enterobactin biosynthesis inhibits the acquisition of microcin E492 post-translational modification, as monitored by high-performance liquid chromatography and mass spectrometry. Furthemore, exogenous enterobactin restored the production of post-translationally modified microcin in a bacterial strain deficient in enterobactin synthesis. We thus concluded that enterobactin serves as a precursor for the synthesis of the post-translationally modified microcin and that the unmodified microcin is an incompletely processed form of the mature microcin E492. Gene disruption experiments showed that MceC and MceD, two enzymes encoded by the mceABCDEFGHIJ gene cluster, are involved in the synthesis of microcin E492 post-translational modification, as followed by mass spectrometry. Genes homologous to iroB and iroD, required for the conversion (linearization and C-glycosylation) of enterobactin into salmochelins, efficiently complemented mceC and mceD, respectively. Based on our results, a model is proposed for the biosynthesis of the mature siderophore-carrying peptide.