Two
membrane transporters, the 17
amino acid (aa)
oligopeptide penetratin derived from the homeodomain of Antennapedia (Ant) and an analogue of the basic domain of TAT (aa 47-57) (TAT-a) from HIV-1, were tested as carriers for a p53 C-terminal
peptide (aa 361-382) into human
breast cancer cells. The studies were performed to determine whether the membrane-transduction efficiency of membrane carriers: Ant, TAT or TAT analogue (TAT-a) correlated with
peptide hydrophobic features.
Peptide-sequence analysis clearly demonstrated that the Ant sequence and p53
peptide sequence (p53p) together created a
peptide with enhanced hydrophobic characteristics; while the TAT or TAT analogue (TAT-a) and p53p sequence together created a
peptide with significantly less hydrophobic qualities. The degree of hydrophobic moment and helical wheel plots for these
peptides correlated directly with their ability to transduce the p53
peptide. Western blot analysis revealed that Ant was able to transduce p53 C-terminal
peptide into human
breast cancer cells as a highly efficient
membrane transporter. Compared to Ant, TAT-a fused to the C-terminus of p53
peptide (p53p-TAT-a) was a less efficient carrier into these cells under the conditions of our study. Additionally, N-terminal linked TAT-a to p53p (TAT-a-p53p) showed even lower efficiency as a transporter than p53-TAT-a. Apoptosis assays showed that the p53
peptide, fused at its C-terminus to Ant (p53p-Ant), induced a higher percentage of apoptotic cells in human
breast cancer cell lines expressing mutant or wild-type p53 as compared to p53
peptide fused at its C-terminus to the TAT-a sequence (p53p-TAT-a) or when fused at the N-terminus to TAT-a (TAT-a-p53p). These data suggested a direct correlation between hydrophobic characteristics and efficiency as a transporter. Sequence study, using hydrophobic moment and helical wheel analyses, may be useful predictive tools for choosing the best carrier for a
peptide.