Cellular Internalization of Quantum Dots Noncovalently Conjugated with Arginine-Rich Cell-Penetrating Peptides

Abstract

Protein transduction domains comprised of basic amino acid-rich peptides, can efficiently deliver covalently fused macromolecules into cells. Quantum dots (QDs) are luminescent semiconductor nanocrystals that are finding increasing application in biological imaging. Previous studies showed that protein transduction domains mediate the internalization of covalently attached QDs. In this study, we demonstrate that arginine-rich intracellular delivery peptides (cell-penetrating peptides; CPPs), analogs of naturally-occuring protein transduction domains, deliver noncovalently associated QDs into living cells; CPPs dramatically increase the rate and efficiency of cellular uptake of QD probes. The optimal molecular ratio between arginine-rich CPPs and QD cargoes for cellular internalization is approximately 60:1. Upon entry into cells, the QDs are concentrated in the perinuclear region. There is no cytotoxicity following transport of QDs present at concentrations up to 200 nM. The mechanism for arginine-rich CPP/QD complexes to traverse cell membrane appears to involve a combination of internalization pathways. These results provide insight into the mechanism of arginine-rich CPP delivery of noncovalently attached cargoes, and may provide a powerful tool for imaging in vivo.

Intracellular delivery of quantum dots mediated by a histidine- and arginine-rich HR9 cell-penetrating peptide through the direct membrane translocation mechanism

“…However, QDs alone do not readily enter cells, and aggregation often occurs after internalization [19,20]. To overcome these limitations, surface modified QDs functionalized by covalent [21e23] or noncovalent [15,24,25] linkages with CPPs, denoted as CPP-QDs or CPP/QDs respectively, have been introduced recently.…”

Applications of Fluorescent Quantum Dots for Reproductive Medicine and Disease Detection

“…The intracellular localization of selective biomolecules for targeting presents additional challenges associated with QDs conjugates delivery within the cells. Available methods for QDs delivery are composed of, but not limited to, positively charged peptides or cell-penetrating peptides on QDs, microinjection, electroporation, or nonspecific or receptor-mediated endocytosis [57, [67][68][69]. Electroporation technique has shown robust and highly efficient delivery of both monomer and aggregate QDs to the cells due to induced electrical pulses that temporarily permeabilize the plasma membrane [70].…”

Cell-penetrating peptides for nanomedicine – how to choose the right peptide

“…In a study by Liu et al CPPs did not need to be covalently associated to QDs to achieve efficient cellular internalization. This non-covalent attachment of histidineand arginine-rich peptides to carboxylated QDs allowed for direct membrane translocation and diminished the setback of endosomal, as well as lysosomal trapping [87]. Equipping QDs with cysteine-terminated NLS peptides, such as the adenovirus-derived CGGFSTSLRARKA, the SV40 large T antigen-derived CGGGPKKKRKVGG or the HIV-1 Tat protein-derived CGGRKKRRQRRRAP by means of the heterobifunctional linker (Succinimidyl-4-[N-maleimidomethyl] cyclohexane-1-carboxylate) led to efficient cellular internalization.…”