RESUMEN
We identify specific acylphosphatase (AcP) residues that interact with silica nanoparticles (SNPs) using a combined NMR spectroscopy and proteomics-mass spectrometry approach. AcP associated with 4- and 15-nm diameter SNPs through a common and specific interaction surface formed by amino acids from the two α-helices of the protein. Greater retention of native protein structure was obtained on 4-nm SNPs than on 15-nm particles, presumably due to greater surface curvature-induced protein stabilization with the smaller SNPs. These results demonstrate that proteins may undergo specific and size-dependent orientation on nanoparticle surfaces. Our approach can be broadly applied to various protein-material systems to help understand in much greater detail the protein-nanomaterial interface; it would also encourage better modeling, and thus prediction and design, of the behavior of functional proteins adsorbed onto different surfaces.
Asunto(s)
Nanopartículas/química , Proteínas/química , Dióxido de Silicio/química , Propiedades de SuperficieRESUMEN
We describe a method for determining the orientation of cytochrome c, RNase A, and lysozyme on silica nanoparticles (SNPs) using chemical modification combined with proteolysis-mass spectrometry. The proteins interacted with SNPs through preferential adsorption sites, which are dependent on SNP diameter; 4 nm SNPs induce greater structural stabilization than 15 nm particles, presumably due to greater surface curvature of the former. These results suggest that nanoparticle size and protein structure influence protein orientation on SNPs.
Asunto(s)
Nanoestructuras/química , Mapeo de Interacción de Proteínas/métodos , Proteínas/química , Dióxido de Silicio/química , Adsorción , Sitios de Unión , Espectrometría de Masas , Ensayo de Materiales , Nanoestructuras/ultraestructura , Unión Proteica , Propiedades de SuperficieRESUMEN
Great strides are being made worldwide in our ability to synthesize and assemble nanoscale building blocks to create advanced materials with novel properties and functionalities. The novel properties of nanostructures are derived from their confined sizes and their very large surface-to-volume ratios. Nanostructured surfaces have also been shown to elicit more favorable and selective biomolecule and cellular responses than surfaces at coarser length scales. In the case of nanoscale ceramics and osteoblasts, for example, the benefit results from protein (vitronectin) unfolding at the nanostructured surface. These nanoscale attributes are enabling a variety of nanostructures to form the bases for a new field--nanomedicine. A fundamental issue in much of nanomedicine, and especially tissue regeneration, is to understand and to eventually control nanostructure-biomolecule interactions. To elucidate the fundamental bases for changes of protein conformation and function on nanostructured surfaces, and hence select responses including those of stem cells, a number of model experiments have been carried out. The results of these studies are presented and discussed in the context of the fundamental driving forces for protein conformation changes associated with nanostructures, their relationship to modified cell responses and tissue engineering, and our present knowledge regarding nanostructure properties.
Asunto(s)
Nanomedicina/métodos , Nanoestructuras/química , Proteínas/metabolismo , Ingeniería de Tejidos/métodos , Nanoestructuras/ultraestructuraRESUMEN
The structure, thermodynamic and kinetic stability, and activity of cytochrome c (cyt c) on silica nanoparticles (SNPs) of different sizes have been studied. Adsorption of cyt c onto larger SNPs results in both greater disruption of the cyt c global structure and more significant changes of the local heme microenvironment than upon adsorption onto smaller SNPs. The disruption of the heme microenvironment leads to a more solvent-accessible protein active site, as suggested by Soret circular dichroism spectroscopy and through an increase in peroxidase activity as a function of increased SNP size. Similarly, the stability of cyt c decreases more dramatically upon adsorption onto larger SNPs. These results are consistent with changes in protein-nanoparticle interactions that depend on the size or surface curvature of the supporting nanostructure. This study provides further fundamental insights into the effects of nanoscale surfaces on adsorbed protein structure and function.
Asunto(s)
Citocromos c/química , Nanopartículas , Dióxido de Silicio , Adsorción , Estabilidad de Enzimas , Tamaño de la Partícula , Conformación ProteicaRESUMEN
This paper reports on the unfolding behavior of ribonuclease A (RNase A) on silica nanoparticle surfaces and quantitively demonstrates that nanoscale size and surface curvature play key roles in influencing the stability of adsorbed proteins. Urea denaturation analyses showed that the thermodynamic stability of RNase A decreased upon adsorption onto the nanoparticles, with greater decrease on larger nanoparticles. The stability changes of RNase A correlate well with the changes in the protein-nanoparticle interactions, which increase as the surface contact area and surface charge interaction increases. This study, therefore, provides fundamental information on the effect of nanoscale surfaces on protein structure and function.