Engineered Recognition of Tetravalent Zirconium and Thorium by Chelator–Protein Systems: Toward Flexible Radiotherapy and Imaging Platforms

Targeted α therapy holds tremendous potential as a cancer treatment: it offers the possibility of delivering a highly cytotoxic dose to targeted cells while minimizing damage to surrounding healthy tissue. The metallic α-generating radioisotopes ²²⁵Ac and ²²⁷Th are promising radionuclides for therapeutic use, provided adequate chelation and targeting. Here we demonstrate a new chelating platform composed of a multidentate high-affinity oxygen-donating ligand 3,4,3-LI­(CAM) bound to the mammalian protein siderocalin. Respective stability constants log β₁₁₀ = 29.65 ± 0.65, 57.26 ± 0.20, and 47.71 ± 0.08, determined for the Eu^III (a lanthanide surrogate for Ac^III), Zr^IV, and Th^IV complexes of 3,4,3-LI­(CAM) through spectrophotometric titrations, reveal this ligand to be one of the most powerful chelators for both trivalent and tetravalent metal ions at physiological pH. The resulting metal–ligand complexes are also recognized with extremely high affinity by the siderophore-binding protein siderocalin, with dissociation constants below 40 nM and tight electrostatic interactions, as evidenced by X-ray structures of the protein:ligand:metal adducts with Zr^IV and Th^IV. Finally, differences in biodistribution profiles between free and siderocalin-bound ²³⁸Pu^IV-3,4,3-LI­(CAM) complexes confirm <i>in vivo</i> stability of the protein construct. The siderocalin:3,4,3-LI­(CAM) assembly can therefore serve as a “lock” to consolidate binding to the therapeutic ²²⁵Ac and ²²⁷Th isotopes or to the positron emission tomography emitter ⁸⁹Zr, independent of metal valence state

The Molecular Basis for Binding of an Electron Transfer Protein to a Metal Oxide Surface

Achieving fast electron transfer between a material and protein is a long-standing challenge confronting applications in bioelectronics, bioelectrocatalysis, and optobioelectronics. Interestingly, naturally occurring extracellular electron transfer proteins bind to and reduce metal oxides fast enough to enable cell growth, and thus could offer insight into solving this coupling problem. While structures of several extracellular electron transfer proteins are known, an understanding of how these proteins bind to their metal oxide substrates has remained elusive because this abiotic–biotic interface is inaccessible to traditional structural methods. Here, we use advanced footprinting techniques to investigate binding between the <i>Shewanella oneidensis</i> MR-1 extracellular electron transfer protein MtrF and one of its substrates, α-Fe₂O₃ nanoparticles, at the molecular level. We find that MtrF binds α-Fe₂O₃ specifically, but not tightly. Nanoparticle binding does not induce significant conformational changes in MtrF, but instead protects specific residues on the face of MtrF likely to be involved in electron transfer. Surprisingly, these residues are separated in primary sequence, but cluster into a small 3D putative binding site. This binding site is located near a local pocket of positive charge that is complementary to the negatively charged α-Fe₂O₃ surface, and mutational analysis indicates that electrostatic interactions in this 3D pocket modulate MtrF–nanoparticle binding. Strikingly, these results show that binding of MtrF to α-Fe₂O₃ follows a strategy to connect proteins to materials that resembles the binding between donor–acceptor electron transfer proteins. Thus, by developing a new methodology to probe protein–nanoparticle binding at the molecular level, this work reveals one of nature’s strategies for achieving fast, efficient electron transfer between proteins and materials