ABSTRACT
In biological systems, chemical and physical transformations of engineered silver nanomaterials (AgENMs) are mediated, in part, by proteins and other biomolecules. Metalloprotein interactions with AgENMs are also central in understanding toxicity and antimicrobial and resistance mechanisms. Despite their readily available thiolate and amine ligands, zinc finger (ZF) peptides have thus far escaped study in reaction with AgENMs and their Ag(I) oxidative dissolution product. We report spectroscopic studies that characterize AgENM and Ag(I) interactions with two ZF peptides that differ in sequence, but not in metal binding ligands: the ZF consensus peptide CP-CCHC and the C-terminal zinc finger domain of HIV-1 nucleocapsid protein p7 (NCp7_C). Both ZF peptides catalyze AgENM (10 and 40 nm, citrate coated) dissolution and agglomeration, two important AgENM transformations that impact bioreactivity. AgENMs and their oxidative dissolution product, Ag(I)(aq), mediate changes to ZF peptide structure and metalation as well. Spectroscopic titrations of Ag(I) into apo-ZF peptides show an Ag(I)-thiolate charge transfer band, indicative of Ag(I)-ZF binding. Fluorescence studies of the Zn(II)-NCp_7 complex indicate that the Ag(I) also effectively competes with the Zn(II) to drive Zn(II) displacement from the ZFs. Upon interaction with AgENMs, Zn(II) bound ZF peptides show a secondary structural change in circular dichroism spectroscopy toward an apo-like structure. The results suggest that Ag(I) and AgENMs may alter ZF protein function within the cell.
ABSTRACT
BACKGROUND: In a biological system, an engineered nanomaterial (ENM) surface is altered by adsorbed proteins that modify ENM fate and toxicity. Thus far, protein corona characterizations have focused on protein adsorption, interaction strength, and downstream impacts on cell interactions. Given previous reports of Ag ENM disruption of Cu trafficking, this study focuses on Ag ENM interactions with a model Cu metalloprotein, Cu(II) azurin. The study provides evidence of otherwise overlooked ENM-protein chemical reactivity within the corona: redox activity. RESULTS: Citrate-coated Ag ENMs of various sizes (10-40 nm) reacted with Cu(II) azurin resulted in an order of magnitude more dissolved ionic silver (Ag(I)(aq)) than samples of Ag ENMs only, ENMs mixed Cu(II) ions, or control proteins such as cytochrome c and horse radish peroxidase. This dramatic increase in ENM oxidative dissolution was observed even when Cu(II) azurin was combined with a diverse mixture of Escherchia coli proteins to mimic the complexity of the cellular conona. SDS PAGE results confirm that the multiprotein ENM corona includes azurin. A Cu(I)(aq) colorimetric indicator confirms Cu(II) azurin reduction upon interaction with Ag ENMs, but not with the addition of ionic silver, Ag(I)(aq). CONCLUSIONS: Cu(II) azurin and 10-40 nm Ag ENMs react to catalyze Ag ENM oxidative dissolution and reduction of the model Cu metalloprotein. Results push the current evaluation of protein-ENM characterization beyond passive binding interactions and enable the proposal of a mechanism for reactivity between a model Cu metalloprotein and Ag ENMs.
