The antibiotic and DNA‐transfecting peptide LAH4 selectively associates with, and disorders, anionic lipids in mixed membranes

Abstract

The histidine-rich amphipathic peptide LAH4 has antibiotic and DNA delivery capabilities. The peptide has a strong affinity for anionic lipids found in the outer membrane of bacterial membranes. A role for anionic lipids in release of cationic plasmid-containing complexes has been proposed previously, and disruption of membrane asymmetry and presentation of phosphatidylserine (PS) in the membrane outer leaflet is a general feature observed in diseased mammalian cells. Therefore, to understand the peptide-lipid interactions in more detail, solid-state NMR experiments on model membranes have been performed. 31P MAS NMR on mixed phosphatidylcholine (PC)/PS and PC/phosphatidylglycerol (PG) membranes has been used to demonstrate a strong interaction between LAH4 and anionic lipids. By using deuterated lipids and wide-line 2H NMR when probing lipid chain order, it is demonstrated that LAH4 preferentially interacts with PS over PC and effectively disorders the anionic PS lipid fatty acyl chains. In addition, we demonstrate that the efficiency of gene transfer in vitro to different cell lines is closely related to the degree of disruption of PS acyl chains for four isomers of LAH4. This work suggests a mechanism of selective destabilization by LAH4 of anionic lipids in the membranes of cells during transfection with implications for nucleic acid delivery in vivo.

Molecular Determinants of Vectofusin-1 and Its Derivatives for the Enhancement of Lentivirally Mediated Gene Transfer into Hematopoietic Stem/Progenitor Cells

“…These data underline the fact that these two functions likely rely on different critical molecular determinants. Cell Line Transfection with Vectofusin-1 Derivatives-LAH4 derivatives have been described previously as nucleic acid transfection agents (31)(32)(33)43). Therefore, we tested the capacity of Vectofusin-1 and some derivatives to transfect 293T cells with an eGFP-expressing plasmid.…”

Incorporation of 2,3‐Diaminopropionic Acid into Linear Cationic Amphipathic Peptides Produces pH‐Sensitive Vectors

“…[4,13,24] The ability of LADap to mediate delivery of the reporter gene luciferase to MRC5-V2 cells was assessed by means of a luciferase based transfection assay (Fig. 3).…”

“…3A), indeed expression mediated by LADap was 246 times greater than that mediated by LAK (p < 0.05). We then compared the transfection efficiency of LADap with that of LAH, LAH4-L1, the most efficient vector peptide from our previous studies [13] , and linear polyethylenimine (L-PEI), another cationic non-viral vector that is available commercially (Fig. 3B).…”

“…[3,4] Histidine rich peptides, as with other polyamines with high buffering capacities between pH 5.1 and pH 7.4, [5][6][7][8][9][10][11][12] are capable of acting as a proton sponge while the membrane disturbing behavior and nucleic acid binding ability of the peptide are also dramatically altered. [13,14] As a result of including the pH sensitive histidines, large numbers of membrane disturbing peptides are released from the nucleic acid-peptide complex during endosomal acidification enhancing the ability of the complex to escape the endosome and reach the cell cytosol and, eventually, nucleus. [14] Although structurally related to its cationic analogues such as lysine and ornithine, 2,3-diaminopropionic acid (Dap) in its free acid form is characterized by distinctly lower proton ionization constants for its two amino groups.…”

Incorporation of a Nuclear Localization Signal in pH Responsive LAH4-L1 Peptide Enhances Transfection and Nuclear Uptake of Plasmid DNA

“…22,23 Previously, our group and others have reported the use of LAH4-L1 peptide for DNA and siRNA delivery. [24][25][26][27][28] LAH4-L1 is a histidine-rich, cationic amphipathic peptide with pH responsive properties. At pH 7, the LAH4-L1 binds with and condenses DNA to form nano-sized complexes which enter cells through endocytosis.…”