Oligomer Formation Propensities of Dimeric Bundle Peptides Correlate with Cell Penetration Abilities

Abstract

LK-3, an amphipathic dimeric peptide linked by two disulfide bonds, and related isomeric bundles were synthesized, and their cell penetrating abilities were investigated. The measurements using size exclusion chromatography and dynamic light scattering show that LK-3 and its isomers form cell penetrating oligomers. Calculations, performed for various types of peptide isomers, elucidate a strong correlation between the amphipathic character of dimers and cell penetration ability. The results suggest that the amphipathicities of LK-3 and related bundle dimers are responsible for their oligomerization propensities which in turn determine their cell penetrating abilities. The observations made in this study provide detailed information about the mechanism of cell uptake of LK-3 and suggest a plausible insight of the early stage of nanoparticle formation of the cell penetrating amphipathic peptides.

Design and Optimization of an α-Helical Bundle Dimer Cell-Penetrating Peptide for In Vivo Drug Delivery

“…Next, the cell-penetrating abilities of the dimers were determined with fluorescence-labeled dimeric bundle peptides using flow cytometry analysis. The first-generation CPP, LK-3, penetrates cells with the highest efficacy while the cell permeabilities of its derivatives decrease as their lengths decrease, as shown in the previous report . Although the dimer, LK-3, penetrates cells at a single-digit nanomolar concentration, the proportion of fluorescent-positive cells increases dramatically at higher concentrations.…”

“…Peptide drugs require peptide length optimization, and shorter peptides are better candidate peptides. As shown in previous studies , of LK-3, a bis-disulfide bridged dimer derivative of LKα14, oxidative dimerization via bis-disulfide formation transforms the C-containing monomeric peptides into dimeric bundle CPPs. The fact that dimer instead of multimer formation occurs at these specific sequences is an exceptional finding because stabilizing intermolecular interactions overcome the entropy factor associated with random oxidative disulfide bond formation.…”

“…Recently, we discovered that a bis-disulfide dimer of the amphipathic α-helical peptide, LK-3 (dimeric bundle of a 16aa-length (LK16) peptide LK-2 with an amino acid sequence LKKLCKLLKKLCKLAG given in Figure 1a), forms stable oligomeric nanoparticles via hydrophobic interactions between dimers, a key characteristic that leads to its high cellpenetrating ability at nanomolar concentrations. 5 This exciting finding stimulated thoughts about the use of this dimeric amphipathic α-helical peptide and its derivatives as pharmaceutically relevant CPPs. In this regard, consideration was given to their toxicities at concentrations used for delivering modulator(s).…”

EJP18 peptide derived from the juxtamembrane domain of epidermal growth factor receptor represents a novel membrane-active cell-penetrating peptide

“…The presence of punctate spots on (or adjacent to) the plasma membrane suggests the occurrence of oligomerisation. It has been noted that peptide dimers of the amphiphilic peptide, LK-3, induced further oligomerisation and the burying of their hydrophobic motifs in the plasma membrane to aid internalisation [50].…”

Stapling of short cell-penetrating peptides for enhanced tumor cell-and-tissue dual-penetration

“…In addition, the increase in peptide hydrophobicity may possibly lead to the formation of nanosized oligomer which was found to be conducive to their cell permeability. 17 However, the hydrophobicity does not always guarantee an enhanced cellular uptake. Other factors may also affect the cellular uptake of the peptides in certain cases, such as the sequences, the chirality, and the cell types.…”