Permeation through Phospholipid Bilayers, Skin‐Cell Penetration, Plasma Stability, and CD Spectra of α‐ and β‐Oligoproline Derivatives

Abstract

After a survey of the special role, which the amino acid proline plays in the chemistry of life, the cell-penetrating properties of polycationic proline-containing peptides are discussed, and the widely unknown discovery by the Giralt group (J. Am. Chem. Soc. 2002, 124, 8876) is acknowledged, according to which fluorescein-labeled tetradecaproline is slowly taken up by rat kidney cells (NRK-49F). Here, we describe details of our previously mentioned (Chem. Biodiversity 2004, 1, 1111) observation that a hexa-β(3)-Pro derivative penetrates fibroblast cells, and we present the results of an extensive investigation of oligo-L- and oligo-D-α-prolines, as well as of oligo-β(2)h- and oligo-β(3)h-prolines without and with fluorescence labels (1-8; Fig. 1). Permeation through protein-free phospholipid bilayers is detected with the nanoFAST biochip technology (Figs. 2-4). This methodology is applied for the first time for quantitative determination of translocation rates of cell-penetrating peptides (CPPs) across lipid bilayers. Cell penetration is observed with mouse (3T3) and human foreskin fibroblasts (HFF; Figs. 5 and 6-8, resp.). The stabilities of oligoprolines in heparin-stabilized human plasma increase with decreasing chain lengths (Figs. 9-11). Time- and solvent-dependent CD spectra of most of the oligoprolines (Figs. 13 and 14) show changes that may be interpreted as arising from aggregation, and broadening of the NMR signals with time confirms this assumption.

Electron Transfer across Helical Peptides

“…The proline amino acid is known to be a secondary‐structure breaker, owing to its inability to form hydrogen bonds with its amine group. Nonetheless, oligoprolines have a unique helical secondary structure, which is different from that of the common α‐helical structure of peptides 104. 105…”

Cell‐Penetrating Peptide Foldamers: Drug‐Delivery Tools

“…b-Oligoarginine was translocated through the cell membrane to am uch greater extent than that of b-oligolysine.T hey also reported that b-oligoprolinef ormed as table secondary structure. [14] The cellular uptake of b-oligoproline was much slower than that of b-oligoarginine. However, b-oligoprolinew as clearly observed in cells by means of fluorescence microscopy,r egardless of whether the peptidec ontained no cationic amino acids.…”

“…β‐Oligoarginine was translocated through the cell membrane to a much greater extent than that of β‐oligolysine. They also reported that β‐oligoproline formed a stable secondary structure . The cellular uptake of β‐oligoproline was much slower than that of β‐oligoarginine.…”

De Novo Design of Cell‐Penetrating Foldamers

“…Pro-containing CPPs tend to form rigid secondary structures, which depend on their peptide sequences, and demonstrate efficient cellular uptake. [32][33][34][35] The most common structure of Pro-containing CPPs is a polyproline II (PPII) helix, which is constructed from oligoproline molecules [(Pro) n ]. [36,37] In fact, (Pro) n -based CPPs with cationic or cationic/hydrophobic sidechain groups form PPII helices and exhibit efficient cellular uptake.…”