Polyarginine enters cells more efficiently than other polycationic homopolymers

TLDR

Cationic homopolymers can cross cell membranes and are used to deliver biopolymers and small molecules. The study examined whether polycationic charge or other structural features underlie cellular uptake by assaying various homopolymers with microscopy and flow cytometry. Cellular entry was quantified using confocal microscopy and flow cytometry across different homopolymers. Arginine polymers of six or more residues entered cells far more efficiently than lysine, ornithine, or histidine polymers of the same length, while shorter peptides were ineffective; altering the arginine side‑chain length had little effect, citrulline polymers did not enter, and uptake was energy‑dependent but not endocytosis‑mediated.

Abstract

Abstract: Homopolymers or peptides containing a high percentage of cationic amino acids have been shown to have a unique ability to cross the plasma membrane of cells, and consequently have been used to facilitate the uptake of a variety of biopolymers and small molecules. To investigate whether the polycationic character of these molecules, or some other structural feature, was the molecular basis for the effect, the ability of a variety of homopolymers to enter cells was assayed by confocal microscopy and flow cytometry. Polymers of l ‐ or d ‐arginine containing six or more amino acids entered cells far more effectively than polymers of equal length composed of lysine, ornithine and histidine. Peptides of fewer than six amino acids were ineffective. The length of the arginine side‐chain could be varied without significant loss of activity. These data combined with the inability of polymers of citrulline to enter cells demonstrated that the guanidine headgroup of arginine was the critical structural component responsible for the biological activity. Cellular uptake could be inhibited by pre‐incubation of the cells with sodium azide, but not by low temperature (3 °C), indicating that the process was energy dependent, but did not involve endocytosis.