H-Bonding-mediated binding and charge reorganization of proteins on gold nanoparticles.

Brahmaiah Meesaragandla,Isabel García,Luis M Liz-Marzán,Ivan Coluzza,Mihaela Delcea,Doreen Biedenweg,Jhoan Toro-Mendoza

doi:10.1039/c9cp06371d

Abstract

Once introduced into the human body, nanoparticles often interact with blood proteins, which in turn undergo structural changes upon adsorption. Although protein corona formation is a widely studied phenomenon, the structure of proteins adsorbed on nanoparticles is far less understood. We propose a model to describe the interaction between human serum albumin (HSA) and nanoparticles (NPs) with arbitrary coatings. Our model takes into account the competition between protonated and unprotonated polymer ends and the curvature of the NPs. To this end, we explored the effects of surface ligands (citrate, PEG-OMe, PEG-NH2, PEG-COOH, and glycan) on gold nanoparticles (AuNPs) and the pH of the medium on structural changes in the most abundant protein in blood plasma (HSA), as well as the impact of such changes on cytotoxicity and cellular uptake. We observed a counterintuitive effect on the ζ-potential upon binding of negatively charged HSA, while circular dichroism spectroscopy at various pH values showed an unexpected pattern in the reduction of α-helix content, as a function of surface chemistry and curvature. Our model qualitatively reproduces the decrease in α-helix content, thereby offering a rationale based on particle curvature. The simulations quantitatively reproduce the charge inversion measured experimentally through the ζ-potential of the AuNPs in the presence of HSA. Finally, we found that AuNPs with adsorbed HSA display lower toxicity and slower cell uptake rates, compared to functionalized systems in the absence of protein. Our study allows examining and explaining the conformational dynamics of blood proteins triggered by NPs and corona formation, thereby opening new avenues toward designing safer NPs for drug delivery and nanomedical applications.

Highlights

The use of gold nanoparticles (AuNPs) in biomedical applications such as drug delivery, cellular targeting, imaging, photodynamic therapy, and tissue engineering[1,2,3,4,5] results from the unique combination of their distinctive physicochemical and optical properties, i.e. low toxicity, biodegradability and localized surface plasmon resonances.[6,7,8] AuNPs can readily form stable conjugates with proteins through either covalent bonds or physical interactions.Upon entering the human body, NPs may interact with blood proteins, forming a so-called ‘‘protein corona’’,9 which governs the ultimate fate of the biological activity of the NPs.[10,11,12,13,14] The formation of a corona can, lead to changes in the structures of the adsorbed proteins, thereby affecting their physiological functions and potentially inducing unexpected biological reactions such as immunostimulation or immunosuppression.[15,16]In particular, various properties of AuNPs such as size, shape and surface chemistry have been reported to influence the binding of proteins to nanoparticles, in terms of protein structure and flexibility, which, in turn, influence how nanoparticles interact with cells and/or tissues.[17,18] The interplay between all such properties has been investigated by numerical simulations[19,20] and, experimentally, by the application of techniques such as circular dichroism (CD) spectroscopy and sum frequency generation vibrational spectroscopy.[21,22] The analysis pursues the effects of the curvature, morphology and chemical nature of the NPs on the conformational changes of proteins under given pH and temperature conditions
Citrate-capped AuNPs showed a localized surface plasmon resonance (LSPR) band at 520 nm, which was preserved after all different surface modifications (Fig. S1, Electronic supplementary information (ESI)†)
The counterintuitive results found in the increased positive charge upon binding of negatively charged human serum albumin (HSA) and the effects of the complex interplay between surface chemistry and NP size on protein conformational changes were successfully explained

Summary

Introduction

The use of gold nanoparticles (AuNPs) in biomedical applications such as drug delivery, cellular targeting, imaging, photodynamic therapy, and tissue engineering[1,2,3,4,5] results from the unique combination of their distinctive physicochemical and optical properties, i.e. low toxicity, biodegradability and localized surface plasmon resonances.[6,7,8] AuNPs can readily form stable conjugates with proteins through either covalent bonds or physical interactions.Upon entering the human body, NPs may interact with blood proteins, forming a so-called ‘‘protein corona’’,9 which governs the ultimate fate of the biological activity of the NPs.[10,11,12,13,14] The formation of a corona can, lead to changes in the structures of the adsorbed proteins, thereby affecting their physiological functions and potentially inducing unexpected biological reactions such as immunostimulation or immunosuppression.[15,16]In particular, various properties of AuNPs such as size, shape and surface chemistry have been reported to influence the binding of proteins to nanoparticles, in terms of protein structure and flexibility, which, in turn, influence how nanoparticles interact with cells and/or tissues.[17,18] The interplay between all such properties has been investigated by numerical simulations[19,20] and, experimentally, by the application of techniques such as circular dichroism (CD) spectroscopy and sum frequency generation vibrational spectroscopy.[21,22] The analysis pursues the effects of the curvature, morphology and chemical nature of the NPs on the conformational changes of proteins under given pH and temperature conditions. For PEG-OMe, PEG-COOH, PEG-NH2, and glycan functionalized AuNPs, the hydrodynamic diameter of the bioconjugates did not increase significantly (Fig. S5, ESI†), likely due to the shielding effect of both PEG and glycans, or to an insufficient amount (2.4 Â 10À6 M) of HSA to form a protein corona.

Results

Conclusion