Cytosolic Diffusion and Peptide-Assisted Nuclear Shuttling of Peptide-Substituted Circa 102 Gold Atom Nanoclusters in Living Cells

Abstract

For biological and medical application, a definable nanomaterial and knowledge on its fate after administration are highly recommended if not mandatory. Here, we synthesized a water-soluble gold nanocluster by sodium borohydride reduction of chloroauric acid in presence of 5,5'-dithio-bis(2-nitrobenzoic acid). The resulting gold nanocluster displayed the physical characteristics of a cluster containing an inner core of about 100 gold atoms that was surrounded by an organic monolayer made of about 30 thioaminobenzoic acids (TAB) and 14 anionic thionitrobenzoic acids (TNB). The mixed TAB-,TNB-protected gold nanocluster reacted well in water with thiolated peptides containing a Nuclear Localization Signal (NLS) or a Nuclear Export Signal (NES) mainly by exchange of the TNB, providing gold nanoclusters equipped with 8-9 intracellular active peptides and a remaining ligand coverage consisting mostly of the zwitterion TAB. The behavior of these peptide-gold nanoclusters inside the cytosol and nucleus of cells was then assayed using an electroporation procedure allowing transient plasma membrane permeability. Light and electron microscopy observations demonstrated a consistent inflow and diffusion of the gold nanoclusters into the cytosol. Inside the living cells, the distribution of the gold nanoparticles was specifically driven by the appended signal peptides in a manner similar to the distribution of NLS and NES-bearing proteins, demonstrating diffusion ability, stability and usage of these definable ligand-substituted gold nanoclusters for intracellular applications.

Ultrasmall Gold Nanoparticles Coated with Zwitterionic Glutathione Monoethyl Ester: A Model Platform for the Incorporation of Functional Peptides

“…The general antifouling properties of ultrasmall AuNPs raise the possibility that they could be conjugated with additional functional moieties providing desired molecular-recognition properties. − Indeed, recent studies have demonstrated the fabrication of ultrasmall AuNPs covered with targeted peptides for the recognition of intracellular proteins and membrane receptors. ,− Nonetheless, additional work is still needed to characterize and understand in more detail the behavior of functional ultrasmall AuNPs in complex media. For example, there is surprisingly little work done on the functionalization of the widely used ultrasmall AuGSH particles with peptides and systematic evaluation of the corresponding constructs in biological media. , …”

Bioactivated and PEG‐Protected Circa 2 nm Gold Nanoparticles for in Cell Labelling and Cryo‐Electron Microscopy

“…The cytotoxicity and distribution of the AuNPs within living cells was investigated after having delivered the probe using a com-mercially available electroporation device [39] and electric pulses that were previously optimized for antibody transduction [15,40] (Figure 3 and Figure 4). AuNPs at the indicated final concentrations were incubated with freshly suspended HeLa cells in PBS.…”

Active U11 Peptide Luminescent Gold Nanoclusters for Pancreatic Tumor Cell Targeting

“…However, passive-targeting AuNCs invariably suffer from low tumor specificity and associated cytotoxic effects, and the introduction of active tumor-targeting elements requires post-synthetic ligand modification. 12,13 The in situ generation of Au NCs inside the cell was successfully used to image the nucleolus of the cell. 9 However, it remains challenging to label the nuclei of the cells with exogenous AuNCs.…”

“…Among these, small gold nanoclusters (AuNCs) have attracted prime attention due to their ultrasmall size, good biocompatibility, and high stability. − Interestingly they can invade cancer cells and reside throughout the cytoplasm, whereas they hardly enter noncancerous cells and preferentially localize at the membrane region of the cell. However, passive-targeting AuNCs invariably suffer from low tumor specificity and associated cytotoxic effects, and the introduction of active tumor-targeting elements requires post-synthetic ligand modification. , The in situ generation of Au NCs inside the cell was successfully used to image the nucleolus of the cell . However, it remains challenging to label the nuclei of the cells with exogenous AuNCs.…”