First direct evidence for direct cell-membrane penetrations of polycationic homopoly(amino acid)s produced by bacteria

Abstract

Bacteria produce polycationic homopoly(amino acid)s, which are characterized by isopeptide backbones. Although the biological significance of polycationic homopoly(amino acid)s remains unclear, increasing attention has recently been focused on their potential use to achieve cellular internalization. Here, for the first time, we provide direct evidence that two representative bacterial polycationic isopeptides, ε-poly-l-α-lysine (ε-PαL) and ε-oligo-l-β-lysine (ε-OβL), were internalized into mammalian cells by direct cell-membrane penetration and then diffused throughout the cytosol. In this study, we used clickable ε-PαL and ε-OβL derivatives carrying a C-terminal azide group, which were enzymatically produced and then conjugated with a fluorescent dye to analyze subcellular localization. Interestingly, fluorescent proteins conjugated with the clickable ε-PαL or ε-OβL were also internalized into cells and diffused throughout the cytosol. Notably, a Cre recombinase conjugate with ε-PαL entered cells and mediated the Cre/loxP recombination, and ε-PαL was found to deliver a full-length IgG antibody to the cytosol and nucleus.

Characterization of PGua4, a Guanidinium-Rich Peptoid that Delivers IgGs to the Cytosol via Macropinocytosis

“…Surprisingly, we also showed that the covalent coupling between PGua 4 and IgGs allowed IgG internalization in endosomes but did not support IgG delivery into the cytosol. This result was unexpected given that most Arg-rich CPPs, such as Tat and oligo-Arg, have been shown to deliver their cargo when covalently linked to them. − This suggests that either covalent binding altered the endosomal escape properties of PGua 4 , as previously reported for other CPPs, or that the ratio of IgGs-PGua 4 was not optimal . Indeed, four PGua 4 peptoids were covalently linked to each IgG, whereas many more PGua 4 peptoids may be complexed to each IgG using the noncovalent complexation method.…”

History of the streptothricin antibiotics and evidence for the neglect of the streptothricin resistome

“…They proposed that the higher toxicity of streptothricins A-E and X compared to streptothricin F may be due to greater host cellular uptake as a result of their longer β-lysine homopolymer tails (Fig. 2a) 65 . A result of this difference in cytotoxicity is that pure streptothricin F, as opposed to a mix of streptothricins, may have a wide enough therapeutic window to justify its use against multidrug-resistant pathogens.…”

Interfacing Coacervates with Membranes: From Artificial Organelles and Hybrid Protocells to Intracellular Delivery

“…Moreover, these cationic bacterial polymers could be conjugated to large antibodies (IgG, 150 kDa), which were directly delivered into the cytosol and nucleus. [82] Besides cell-penetrating peptides and polymers, various nanoparticles have also been shown to be capable of membrane penetration. [77] Löwen and co-workers developed a simple model to describe the interaction of active particles with minimal membranes, which allows for both penetrations of the membrane (with and without healing) and membrane trapping (endocytosis) (Figure 7e-g).…”

“…Moreover, these cationic bacterial polymers could be conjugated to large antibodies (IgG, 150 kDa), which were directly delivered into the cytosol and nucleus. [ 82 ]…”