Ferric ion (hydr)oxo clusters in the "Venus flytrap" cleft of FbpA

Isothermal calorimetric studies of the binding of iron(III) citrate to ferric ion binding protein from Neisseria gonorrhoeae suggested the complexation of a tetranuclear iron(III) cluster as a single step binding event (apparent binding constant K (app) (ITC) = 6.0(5) x 10(5) M-1). High-resolution Fourier transform ion cyclotron resonance mass spectrometric data supported the binding of a tetranuclear oxo(hydroxo) iron(III) cluster of formula [Fe4O2(OH)(4)(H2O)(cit)](+) in the interdomain binding cleft of FbpA. The mutant H9Y-nFbpA showed a twofold increase in the apparent binding constant [K (app) (ITC) = 1.1(7) x 10(6) M-1] for the tetranuclear iron(III) cluster compared to the wild-type protein. Mossbauer spectra of Escherichia coli cells overexpressing FbpA and cultured in the presence of added Fe-57 citrate were indicative of the presence of dinuclear and polynuclear clusters. FbpA therefore appears to have a strong affinity for iron clusters in iron-rich environments, a property which might endow the protein with new biological functions.