Functionalization with C-terminal cysteine enhances transfection efficiency of cell-penetrating peptides through dimer formation
Artikel i vetenskaplig tidskrift, 2012

Cell-penetrating peptides have the ability to stimulate uptake of macromolecular cargo in mammalian cells in a non-toxic manner and therefore hold promise as efficient and well tolerated gene delivery vectors. Non-covalent peptide-DNA complexes (‘‘peptiplexes’’) enter cells via endocytosis, but poor peptiplex stability and endosomal entrapment are considered as main barriers to peptide- mediated delivery. We explore a simple, yet highly efficient, strategy to improve the function of peptide-based vectors, by adding one terminal cysteine residue. This allows the peptide to dimerize by disulfide bond formation, increasing its affinity for nucleic acids by the ‘‘chelate effect’’ and, when the bond is reduced intracellularly, letting the complex dissociate to deliver the nucleic acid. By introducing a single C-terminal cysteine in the classical CPP penetratin and the penetratin analogs PenArg and EB1, we show that this minor modification greatly enhances the transfection capacity for plasmid DNA in HEK293T cells. We conclude that this effect is mainly due to enhanced thermodynamic stability of the peptiplexes as endosome-disruptive chloroquine is still required for transfection and the effect is more pronounced for peptides with lower inherent DNA condensation capacity. Interestingly, for EB1, addition of one cysteine makes the peptide able to mediate transfection in absence of chloroquine, indicating that dimerisation can also improve endosomal escape properties. Further, the cytotoxicity of EB1 peptiplexes is considerably reduced, possibly due to lower concentration of free peptide dimer resulting from its stronger binding to DNA.

Disulfide bond

Peptiplex

Cysteine

Gene delivery

Dimerization

Cell-penetrating peptides

Författare

Helene Åmand

Chalmers, Kemi- och bioteknik, Fysikalisk kemi

Bengt Nordén

Chalmers, Kemi- och bioteknik, Fysikalisk kemi

Kristina Fant

Chalmers, Kemi- och bioteknik, Fysikalisk kemi

Biochemical and Biophysical Research Communications

0006-291X (ISSN) 1090-2104 (eISSN)

Vol. 418 3 469-474

Ämneskategorier

Fysikalisk kemi

Biokemi och molekylärbiologi

Styrkeområden

Energi

Livsvetenskaper och teknik

Materialvetenskap

Fundament

Grundläggande vetenskaper

DOI

10.1016/j.bbrc.2012.01.041