Active transport across the outer membrane in gram-negative bacteria requires the energy that is generated by the proton motive force in the inner membrane. This energy is transduced to the outer membrane by the TonB protein in complex with the proteins ExbB and ExbB. In the pathogen Vibrio anguillarum we have identified two TonB systems, TonB1 and TonB2, the latter is used for ferric-anguibactin transport and is transcribed as part of an operon that consists of orf2, exbB2, exbD2, and tonB2. This cluster was identified by a polar transposon insertion in orf2 that resulted in a strain deficient for ferric-anguibactin transport. Only the entire cluster (orf2, exbB2, exbD2 and tonB2) could complement for ferric-anguibactin transport, while just the exbB2, exbD2, and tonB2 genes were unable to restore transport. This suggests an essential role for this OrO, designated TtpC, in TonB2-mediated transport in V. anguillarum. A similar gene cluster exists in V. cholerae, i.e., with the homologues of ttpC-exbB2-exbD2-tonB2, and we demonstrate that TtpC from V. cholerae also plays a role in the TonB2-mediated transport of enterobactin in this human pathogen. Furthermore, we also show that in V. anguillarum the TtpC protein is found as part of a complex that might also contain the TonB1, ExbB2, and ExbB2 proteins. This novel component of the TonB1 system found in V. anguillarum and V. cholerae is perhaps a general feature in bacteria harboring the Vibrio-like TonB2 system. Copyright © 2007, American Society for Microbiology. All Rights Reserved.