Microcins are gene-encoded antimicrobial peptides secreted by Enterobacteriaceae. Among them, microcin J25 (MccJ25) is naturally produced by Escherichia coli AY25. The 2.1 kDa mature MccJ25 (21 residues) was isolated from culture supernatants. This lasso-type peptide presents a side chain to backbone cyclization between Glu8 and the N-terminus. The embedded ring is threaded by the C-terminal tail of the molecule, thus forming a noose-like feature. Upon thermolysin-cleavage, a two-chain analogue is obtained (t-MccJ25) that retains the lasso-structure of MccJ25 but looses the Val11-Pro16 hairpin-like structure. The cleaved peptide is about 30 to 100-fold less active than MccJ25 against E. coli and Salmonella enterica species. In early studies, bacteria resistant to MccJ25 were reported to display mutations in the multifunctional outer membrane protein FhuA, the high affinity transporter of iron-ferrichrome and the receptor of phages T5, T1 and phi80. In this study, we have investigated the potential role of FhuA as a receptor for MccJ25 and compared the interaction of MccJ25 and t-MccJ25 with FhuA and their respective inhibitory activity on FhuA binding properties.