Iron-free pyoverdin binds to its outer membrane receptor FpvA in Pseudomonas aeruginosa: a new mechanism for membrane iron transport

Under iron limitation, Pseudomonas aeruginosa secretes a fluorescent sidero phore called pyoverdin, which, after complexing iron, is transported back i nto the cell via its outer membrane receptor FpvA. Previous studies demonst rated co-purification of FpvA with iron-free PaA and reported similar bindi ng affinities of iron-free pyoverdin and ferric-pyoverdin to purified FpvA. The fluorescence resonance energy transfer between iron-free PaA and the F pvA receptor here reveals the existence of an FpvA-pyoverdin complex in P. aeruginosa in vivo, suggesting that the pyoverdin-loaded FpvA is the normal state of the receptor in the absence of iron. Using tritiated ferric-pyove rdin, it is shown that iron-free PaA binds to the outer membrane but is not taken up into the cell, and that in vitro and, presumably, in vivo ferric- pyoverdin displaces the bound iron-free pyoverdin on FpvA-PaA to form FpvA- PaA-Fe complexes. In vivo, the kinetics of formation of this FpvA-PaA-Fe co mplex are more than two orders of magnitude faster than in vitro and depend on the presence of TonB. In P. aeruginosa, two tonB genes have been identi fied (tonB1 and tonB2). TonB1 is directly involved in ferric-pyoverdin upta ke, and TonB2 seems to be able partially to replace TonB1 in its role in ir on acquisition. However, no effect of TonB1 or TonB2 on the apparent affini ty of free pyoverdin to FpvA was observed, and a 17-fold difference was mea sured between the affinities of the two forms of pyoverdin (PaA and PaA-Fe) to FpvA in the absence of TonB1 or TonB2. The mechanism of iron uptake in P. aeruginosa via the pyoverdin pathway is discussed in view of these new f indings.