Protein-Protein Stabilization in V^IVO/8-Hydroxyquinoline-Lysozyme Adducts

The binding of the potential drug [V^IVO(8-HQ)₂], where 8-HQ is 8-hydroxyquinolinato, with hen egg white lysozyme (HEWL) was evaluated through spectroscopic (electron paramagnetic resonance, EPR, and UV-visible), spectrometric (electrospray ionization-mass spectrometry, ESI-MS), crystallographic (X-ray diffraction, XRD), and computational (DFT and docking) studies. ESI-MS indicates the interaction of [V^IVO(8-HQ)(H₂O)]⁺ and [V^IVO(8-HQ)₂(H₂O)] species with HEWL. Room temperature EPR spectra suggest both covalent and non-covalent binding of the two different V-containing fragments. XRD analyses confirm these findings, showing that [V^IVO(8-HQ)(H₂O)]⁺ interacts covalently with the solvent exposed Asp119, while cis-[V^IVO(8-HQ)₂(H₂O)] non-covalently with Arg128 and Lys96 from a symmetry mate. The covalent binding of [V^IVO(8-HQ)(H₂O)]⁺ to Asp119 is favored by a π-π contact with Trp62 and a H-bond with Asn103 of a symmetry-related molecule. Additionally, the covalent binding of V^VO₂ ⁺ to Asp48 and non-covalent binding of other V-containing fragments to Arg5, Cys6, and Glu7 are revealed. Molecular docking indicates that, in the absence of the interactions occurring at the protein-protein interface close to Asp119, the covalent binding to Glu35 or Asp52 should be preferred. Such a protein-protein stabilization could be more common than what believed up today, at least in the solid state, and should be considered in the characterization of metal-protein adducts.