Solution NMR of Nanoparticles in Serum: Protein Competition Influences Binding Thermodynamics and Kinetics

Abstract

The spontaneous formation of a protein corona on a nanoparticle surface influences the physiological success or failure of the synthetic nanoparticle as a drug carrier or imaging agent used in vivo. A quantitative understanding of protein-nanoparticle interactions is therefore critical for the development of nanoparticle-based therapeutics. In this perspective, we briefly discuss the challenges and limitations of current approaches used for studying protein-nanoparticle binding in a realistic biological medium. Subsequently, we demonstrate that solution nuclear magnetic resonance (NMR) spectroscopy is a powerful tool to monitor protein competitive binding in a complex serum medium in situ. Importantly, when many serum proteins are competing for a gold nanoparticle (AuNP) surface, solution NMR is able to detect differences in binding thermodynamics, and kinetics of a tagged protein. Combined with other experimental approaches, solution NMR is an invaluable tool to understand protein behavior in the nanoparticle corona.

Understanding the Adsorption of Peptides and Proteins onto PEGylated Gold Nanoparticles

“…For comparison, the binding of these biomolecules onto non-PEGylated AuNPs is extremely fast and results in higher binding capacities. Previous studies show that ~90% of protein binding to citrate-coated AuNPs occurs in the initial ~5 min, and adsorption is completed within an hour [ 38 , 39 ]. The binding capacities (molecules per AuNp) for GSH, WT GB3 and BSA on bare AuNPs were determined to be , , and , respectively [ 37 ].…”

“…The AuNPs are coated with 5K, 10K, and 30K thiolated PEG, allowing us to probe different levels of PEG coverage. Using NMR spectroscopy [ 37 , 38 , 39 ], we have quantified the kinetics of and final stoichiometry of biomolecular adsorption to PEGylated AuNPs, and we discuss trends and physical properties that lead to efficient binding to PEGylated AuNP surfaces.…”

Chemical and Biophysical Signatures of the Protein Corona in Nanomedicine

“…Protein NMR signals, including their peak intensities, chemical shifts, and line widths, are perturbed as proteins interact with nanoparticles. Specifically, solution NMR spectroscopy can monitor the competitive binding of serum proteins for AuNP surface in situ and detect differences in binding thermodynamics for tagged proteins. ,, However, NMR is cost-intensive, which requires uniform isotopic labeling of proteins. Other in silico techniques like MD simulations have been developed to investigate the interactions of proteins with nanoparticles, ,,,,, exploiting their precise identification of the structural components that drive protein–nanoparticle interaction.…”

Predicting protein function and orientation on a gold nanoparticle surface using a residue-based affinity scale

“…Therefore, we generate an experimental affinity scale for each residue by varying a key binding position in the small GB3 protein 18 . Competitive protein binding is required to rationalize residue differences 19,20 , demonstrating that both kinetic and thermodynamic considerations influence adsorption [21][22][23] . This affinity scale and a simple surface-area based algorithm predict the preferred orientation/function for three proteins presented here, including two blind predictions; moreover, our findings rationalize the observed function of five additional proteins from the literature.…”