Promotion of protein assembly by nanoparticles: From in vitro to in-cell.

Abstract

Engineered nanomaterials are increasingly being studied for biomedical applications such as in diagnosis and therapeutics, due to their similarity in size to subcellular scales, and their tunable physico-chemical properties. Nanomaterials acquire a biomolecular corona upon introduction to biological media, which can influence their biological identity and subsequently, their interactions within the cell. Nanoparticle-protein interactions at the nano-bio interface also affect proteins in the corona, leading to biological transformations such as a change in protein function, unmasking new epitopes, and fibrilization. Silica nanoparticles - one of the most widely studied nanomaterials in biomedical applications - were reported to show a dose-dependent or surface ligand-dependent effect on the kinetics and morphology of protein self-assembly in systems like amyloid-β fibrils, α-synuclein and lysozyme. The interaction of silica nanoparticles with polymerizing cytoskeletal proteins, specifically tubulin proteins, has not been investigated quantitatively. BtubA and BtubB are a pair of bacterial tubulin proteins identified in Prosthecobacter strains, acquired by bacteria from eukaryotic cells via horizontal gene transfer. BtubAB dimers are known to self-assemble like eukaryotic tubulin, forming four-stranded microtubules in vitro as well as five-stranded microtubules in vivo. FRET-labeling of each of the Btub monomers with a donor (mEGFP) and acceptor (mRuby3) fluorescent protein provides a quantitative tool to measure the binding interactions of BtubAB dimers in the presence of silica nanoparticles in vitro and in cell using fluorescence spectroscopy and microscopy. We show that silica nanoparticles enhance BtubAB dimerization in vitro, whereas they appear to have little effect on bacterial tubulin self-assembly in the complex mammalian cellular environment.