Role of tripartite efflux pumps in pyoverdine secretion in Pseudomonas putida KT2440

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Iron is essential for most organisms, but often difficult for bacteria to acquire because iron ions have a low solubility in natural habitats or are bound to proteins in hosts. Bacteria of the genus Pseudomonas solve this problem by the synthesis, secretion and re-uptake of the iron scavenging siderophore pyoverdine (PVD). PVD synthesis and uptake of the PVD-iron complex are relatively well investigated, however, the mechanism of PVD secretion is only partially understood. This project explores the mechanism of PVD secretion using the soil bacterium Pseudomonas putida KT2440 as experimental model. The strain is the best studied saprophytic pseudomonad, a functional chassis for industrial biocatalysis, and it produces only one type of siderophore, PVD. Our preliminary results suggest that PVD secretion does not depend on a particular efflux system, but on several so-called tripartite efflux pumps, which are in part known to contribute to drug resistance. We plan (1) to identify the efflux pumps involved in PVD secretion in P. putida KT2440; (2) to investigate the specific role of these pumps in PVD-mediated iron acquisition and in the resistance against drugs and other toxic compounds; and (3) to characterize PVD transport for the first time at the biochemical level to test a direct involvement of tripartite efflux pumps in PVD transport and to obtain information on the mechanism of energy coupling, substrate specificity and kinetics. The results are expected to contribute to a better understanding of the physiology of pseudomonads under iron limitation and to shed new light on the physiological significance of tripartite efflux pumps.