Protein transduction domains (PTDs), such as the third helix of the Drosophila Antennapedia homeobox gene ( Antp) and the HIV TAT PTD, posses a characteristic positive charge on the basis of their enrichment for arginine and lysine residues. To determine whether cationic peptides are able to function as protein transduction domains, 12-mer peptide sequences from an M13 phage library were selected for synthesis on the basis of their varying cationic charge content. In addition, polylysine and polyarginine peptideswere synthesized in order to assessthe effect of charge contribution in protein transduction. Coupling of the biotinylated peptides to avidin-β-galactosidase facilitated transduction in a wide variety of cell lines and primary cells, including islet β-cells, synovial cells, polarized airway epithelial cells, dendritic cells, myoblasts, and tumor cells. Two of the peptides, PTD-4 and PTD-5, mediated transduction nearly 600-fold more efficiently than a random control peptide, but with an efficiency similar to the TAT PTD and the 12 mers of polylysine and polyarginine. Furthermore, confocal analysis of biotinylated peptide-streptavidin-Cy3 conjugates demonstrated that the internalized PTDs are found in both the nuclei and the cytoplasm of treated cells. When tested in vivo, the PTDs were able to facilitate efficient and rapid protein delivery into rabbit synovium and mouse solid tumors following intraarticular and intratumoral administration, respectively. These novel PTDs can be used to transfer therapeutic proteins and DNA for the treatment of a wide variety of diseases, including arthritis and cancer.
Bibliographical noteFunding Information:
The authors thank Helga Georgescu, Seon-Hee Kim, Andrea Gambotto, Nick Giannoukakis, and Joseph Pilewski for providing cell lines, as well as Eric Lechman and Bruce Baldwin for their technical assistance. The authors also thank Simon Watkins and the Structural Biology and Imaging Center for assistance. This work was partially supported by a grant from the Cystic Fibrosis Foundation and by Contract AR-6-2225 from the National Institutes of Health.
- Cationic peptides
- Protein transduction domains