Binding of cationic alpha-helical peptides to plasmid DNA and their gene transfer abilities into cells.
Polycationic reagents such as cationic lipids and poly-L-lysine are widely used for gene transfer into cells in vitro and show promise as vectors for in vivo gene therapy applications as nonviral gene transfer techniques. We have developed a novel transfection method using cationic amphiphilic alpha-helical oligopeptides with repeated sequences. Oligopeptide has the advantages of being easily designed and modified because of its simple structure. In this study, we synthesized five kinds of peptides of which the total chain length and the width of the hydrophobic region were changed. The binding of the peptides to plasmid DNA was evaluated by agarose gel electrophoresis. It was found that the long and/or hydrophobic peptides can strongly bind to the DNA. The formation of large aggregates with a 0.5-5-microm diameter, which consisted of the long peptides and the DNA, was observed by electron microscopy. The transfection abilities of the peptides were determined by the expression of luciferase from its cDNA in COS-7 cells. The long peptides showed high transfection abilities. As a result, it could be said that the transfection ability of these peptides was parallel to their ability to form aggregates with DNA. Furthermore, the transfection ability was increased by the addition of chloroquine in the transfection procedure. This result indicated that the internalization of the peptide-DNA aggregates would be mediated by the endocytosis pathway.
Niidome
T
T
Department of Applied Chemistry, Faculty of Engineering, Nagasaki University, Nagasaki 852, Japan.
Ohmori
N
N
Ichinose
A
A
Wada
A
A
Mihara
H
H
Hirayama
T
T
Aoyagi
H
H
D000818:Animals; D019556:COS Cells; D002942:Circular Dichroism; D004274:DNA, Recombinant / Q000378:metabolism*; D018014:Gene Transfer Techniques; D008854:Microscopy, Electron; D009842:Oligopeptides / Q000737:chemistry / Q000378:metabolism*; D010957:Plasmids; D011487:Protein Conformation; D014162:Transfection
D004274:DNA, Recombinant; D009842:Oligopeptides