W0304

Structural Evidence for Two-step Ligand Recognition in the Ferric Citrate Receptor FecA. Wyatt Yue, Sylvestre Grizot, Susan Buchanan, Dept. of Crystallography, Birkbeck College, Malet St., London, WC1E 7HX UK.

E. coli possesses a TonB-dependent transport system, which exploits the iron-binding capacity of citrate and its bioavailability for iron acquisition. Here we describe three structures of the outer membrane ferric citrate transporter FecA: unliganded and complexed with iron-free or diferric dicitrate. The structure of FecA bound with iron-free dicitrate represents the first structure of a TonB-dependent transporter bound with an iron-free siderophore. We provide the first evidence that FecA recognizes its cognate siderophore in two steps: FecA initially binds iron-free dicitrate, followed by iron extraction or siderophore displacement in or near the binding pocket to form diferric dicitrate. Binding of diferric dicitrate to FecA results in changes in the orientation of the two citrate ions relative to each other and in their interactions with FecA, compared to iron-free dicitrate. The changes in ligand binding are accompanied by conformational changes in three areas of FecA: two extracellular loops, one plug domain loop and the periplasmic TonB-box motif. The positional and conformational changes in the siderophore and transporter initiate two independent events: ferric citrate transport into the periplasm and transcription induction of the fec transport genes.