Metal clusters in a bacterial iron-trafficking protein

• Main Author: Zhu, Haizhong
• Published: University of Edinburgh 2003
• Subjects: 571.4
• Online Access: http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.664238

Bacterial resistance to standard antibiotic therapies is a critical health protein worldwide. Antimicrobial-resistant infections that are essentially untreatable have begun to occur as epidemics around the globe. In this thesis, I have used the techniques of coordination chemistry, molecular biology, and protein x-ray crystallography to study metal binding of ferric ion binding protein (Fbp). Fbp is a highly-conserved 34 kDa periplasmic iron transporter, a candidate as target for novel metalloantibiotic design. I have discovered that the characteristic dityrosyl motif of Fbp from the human pathogen <i>Neisseria gonorrhoeae</i> can assemble/bind metal clusters in the open metal-binding cleft of the protein, although closure of the hinged metal-binding cleft and the presence of the synergistic phosphate anion are usually considered obligatory for strong metal binding by Fbp. I have also observed that the phosphate ion in the crystal structure of <i>apo</i>-Fbp can be displaced by mononuclear [Hf(NTA)₂]^2-, mononuclear [Fe(NTA)₂]^3- and binuclear [Fe₂(cit)₂(H₂O)₂]^2-, yielding a range of oxo Hf(IV) clusters containing from 3 to 5 metal ions, and oxo Fe(III) clusters containing 3 metal ions. Structural comparisons among the crystal structures of <i>apo</i>-Fbp (1.9 A resolution) and the Hf-bound form of Fbp (1.7 A resolution) and Fe-NTA-Fbp (1.8 A resolution) and Fe₃-FBP (1.8 A resolution) show no hinge closure upon metal binding, suggesting a novel metal acquisition mechanism. The binding of vanadate to <i>apo</i>-FBP has been studied by ⁵¹V NMR, and the data suggest that vanadate can bind similarity to phosphate and also as a vanadium cluster. Crystals of V-Fbp were obtained which diffract to 2.0 A. The work reveals the possibility that bacterial metal uptake is mediated by novel Fbp intermediates including cluster adducts.