We designed a minimalistic zinc(II)-binding peptide featuring the Cys2His2 zinc-finger motif. To this aim, several tens of thousands of (His/Cys)-Xn-(His/Cys) protein fragments (n=2-20) were first extracted from the 3D protein structures deposited in Protein Data Bank (PDB). Based on geometrical constraints positioning two Cys (C) and two His (H) side chains at the vertices of a tetrahedron, approximately 22 000 sequences of the (H/C)-Xi-(H/C)-Xj-(H/C)-Xk-(H/C) type, satisfying Nmetal-binding H=Nmetal-binding C=2, were processed. Several other criteria, such as the secondary structure content and predicted fold stability, were then used to select the best candidates. To prove the viability of the computational design experimentally, three peptides were synthesized and subjected to isothermal calorimetry (ITC) measurements to determine the binding constants with Zn2+, including the entropy and enthalpy terms. For the strongest Zn2+ ions binding peptide, P1, the dissociation constant was shown to be in the nanomolar range (KD=~220 nM; corresponding to ΔGbind=-9.1 kcal mol-1). In addition, ITC showed that the [P1 : Zn2+] complex forms in 1 : 1 stoichiometry and two protons are released upon binding, which suggests that the zinc coordination involves both cysteines. NMR experiments also indicated that the structure of the [P1 : Zn2+] complex might be quite similar to the computationally predicted one. In summary, our proof-of-principle study highlights the usefulness of our computational protocol for designing novel metal-binding peptides.

• Klíčová slova
• Computer design, Isothermal calorimetry, Metal-binding peptide, NMR, QM modeling, Zinc(II),
• MeSH
• molekulární modely MeSH
• peptidy * chemie   metabolismus   chemická syntéza   MeSH
• sekvence aminokyselin MeSH
• termodynamika MeSH
• vazba proteinů MeSH
• zinek * chemie   metabolismus   MeSH
• zinkové prsty MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• peptidy * MeSH
• zinek * MeSH

A rationally-designed scaffold of cyclic octapeptides composed of two units of the natural tripeptide glutathione (GSH) was optimized to strongly and selectively capture toxic lead ions (Pb(II)). Using state-of-the-art computational tools, a list of eleven plausible peptides was shortened to five analogs based on their calculated affinity to Pb(II) ions. We then synthesized and investigated them for their abilities to recover Pb-poisoned human cells. A clear pattern was observed from the in vitro detoxification results, indicating the importance of cavity size and polar moieties to enhance metal capturing. These, together with the apparent benefit of cyclizing the peptides, improved the detoxification of the two lead peptides by approximately two folds compared to GSH and the benchmark chelating agents against Pb poisoning. Moreover, the two peptides did not show any toxicity and, therefore, were thoroughly investigated to determine their potential as next-generation remedies for Pb poisoning.

• Klíčová slova
• Bioinorganic chemistry, Chelates, Glutathione, Lead, Peptides,
• MeSH
• antioxidancia MeSH
• chelátory MeSH
• glutathion * MeSH
• lidé MeSH
• olovo * toxicita   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• antioxidancia MeSH
• chelátory MeSH
• glutathion * MeSH
• olovo * MeSH