Nanoparticle-assisted tubulin assembly is environment dependent

Area of Science:

• Biophysics
• Nanomedicine
• Cell Biology

Background:

• Nanomaterials interact with biological systems, forming a biomolecular corona that alters protein behavior.
• Studying nanoparticle effects on protein interactions typically occurs ex vivo in buffer, lacking in vivo cellular context.
• Bacterial tubulin proteins (BtubA/BtubB) from Prosthecobacter self-assemble similarly to eukaryotic tubulin.

Purpose of the Study:

• To quantitatively compare the effect of silica nanoparticles on bacterial tubulin (BtubAB) protein association in vitro versus within live mammalian cells.
• To investigate the role of the protein corona in mediating nanoparticle-protein interactions.
• To assess the translatability of in vitro findings to the cellular environment.

Main Methods:

• Utilized Förster resonance energy transfer (FRET) with mEGFP and mRuby3 fluorescent labels on BtubA and BtubB monomers.
• Measured BtubAB dimerization and self-assembly using fluorescence spectroscopy and microscopy.
• Introduced unfunctionalized silica nanoparticles to BtubAB in both cell-free buffer and mammalian cells.

Main Results:

• Silica nanoparticles significantly enhanced BtubAB dimerization in buffer, attributed to protein corona formation.
• In contrast, these nanoparticles exhibited minimal effect on bacterial tubulin self-assembly within the mammalian cell environment.
• The complex cellular milieu, with competing biomolecules, influences nanoparticle-protein interactions.

Conclusions:

• Nanoparticle-induced protein-protein interactions observed in vitro may not directly translate to cellular environments.
• Protein corona formation in buffer enhances interactions, but this effect is diminished in vivo due to competitive binding.
• Surface functionalization of nanoparticles is crucial for targeted interactions and overcoming competitive binding in complex biological systems.