Protein Nanoparticle Charge and Hydrophobicity Govern Protein Corona and Macrophage Uptake

Abstract

Protein nanoparticles are biomaterials composed entirely of proteins, with the protein sequence and structure determining the nanoparticle physicochemical properties. Upon exposure to physiological or environmental fluids, it is likely that protein nanoparticles, like synthetic nanoparticles, will adsorb proteins and this protein corona will be dependent on the surface properties of the protein nanoparticles. As there is little understanding of this phenomenon for engineered protein nanoparticles, the purpose of this work was to create protein nanoparticles with variable surface hydrophobicity and surface charge and establish the effect of these properties on the mass and composition of the adsorbed corona, using the fetal bovine serum as a model physiological solution. Albumin, cationic albumin, and ovalbumin cross-linked nanoparticles were developed for this investigation and their adsorbed protein coronas were isolated and characterized by gel electrophoresis and nanoliquid chromatography mass spectrometry. Distinct trends in corona mass and composition were identified for protein nanoparticles based on surface charge and surface hydrophobicity. Proteomic analyses revealed unique protein corona patterns and identified distinct proteins that are known to affect nanoparticle clearance in vivo. Further, the protein corona influenced nanoparticle internalization in vitro in a macrophage cell line. Altogether, these results demonstrate the strong effect protein identity and properties have on the corona formed on nanoparticles made from that protein. This work builds the foundation for future study of protein coronas on the wide array of protein nanoparticles used in nanomedicine and environmental applications.

Measuring the Concentration of Protein Nanoparticles Synthesized by Desolvation Method: Comparison of Bradford Assay, BCA Assay, hydrolysis/UV Spectroscopy and Gravimetric Analysis

“…4A,B; Fig. S4, S5), as previously reported (Pustulka, 2020). In the paper by Tarhini et al, the BSA band was detected in PAGE of BSA nanoparticles (Tarhini, 2018).…”

A Tri-Stimuli Responsive (Maghemite/PLGA)/Chitosan Nanostructure with Promising Applications in Lung Cancer

“…No significant effect on size was observed when the ( γ -Fe 2 O 3 /PLGA)/CS NPs were kept in contact with BSA (34, and 54 mg/mL concentrations in PBS), being the particle diameter and PdI values ≈ 340 nm and ≈ 0.335, respectively. That negligible effect could be attributed to the hydrophilic nature of the CS coating which could minimize protein corona formation and particle aggregation [ 46 , 47 ].…”

“…Additionally, CS is a biocompatible and water-soluble polymer that could be considered as an alternative to poly(ethylene glycol) (PEG) chains when engineering long-circulating NPs. Hence, surface functionalization with CS may optimize the biodistribution and therapeutic effects of PLGA-based nanomedicines by: (i) providing hydrophilic and positively charged stealth coatings that could reduce or even inhibit protein corona formation, thus minimizing and delaying the opsonization process to evade phagocytosis [ 44 , 45 , 46 , 47 ], while favouring the uptake by targeted cells [ 48 , 49 ]; and, (ii) creating an additional barrier to drug diffusion during the early-time burst release of the biphasic drug release profile [ 50 , 51 ]. Incorporation of the CS shell onto a PLGA-based NP may take place by an attractive interaction between the negative PLGA matrix and the positive polysaccharide [ 52 ], and the idea has been applied to the production of magnetopolymer particles, i.e., (Fe 3 O 4 /PLGA)/CS [ 53 , 54 ] and ( γ -Fe 2 O 3 /PLGA)/CS [ 55 ] nanostructures.…”

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

“…These approaches are often able to differentiate many proteins, but they are often unable to study nanoparticle binding in situ. Specifically, mass spectrometric and electrophoretic techniques are widely used to quantify multiple proteins attached in the protein corona (Monopoli et al, 2011;Mo et al, 2018;Zhang et al, 2019;Han et al, 2020;Madathiparambil Visalakshan et al, 2020;Moon et al, 2020;Pinals et al, 2020;Liessi et al, 2021) even at the detail of individual protein residues (Pustulka et al, 2020). However, these methods require extensive purification of the protein-nanoparticle complexes and complete displacement of bound proteins from the nanoparticle surface for analysis.…”