CONFORMATIONAL AND TOPOLOGICAL REQUIREMENTS OF CELL-PERMEABLE PEPTIDEFUNCTION

Cell-permeable peptide import recently was developed to deliver synthe tic peptides into living cells for studying intracellular protein func tions. This import process is mediated by an N-terminal carrier sequen ce which is the hydrophobic region of a signal peptide. In this study, the conformational consequence of the interaction of cell-permeable p eptides with different mimetic membrane environments was investigated by circular dichroism analysis. We showed that cell-permeable peptides adopted or-helical structures in sodium dodecyl sulfate (SDS) micelle s or aqueous trifluoroethanol (TFE). The potency of these peptides in forming helical structures is higher in an amphiphilic environment (SD S) than in a hydrophobic environment (TFE), suggesting that some hydro philic molecules associated with the cell membrane may be involved in peptide import. We also studied topological requirements of cell-perme able peptide function. We demonstrated that peptides containing the ca rrier sequence in their C-termini can also be imported into cells effi ciently. This important discovery can avoid repetitious synthesis of t he membrane-translocating sequence for peptides with different functio nal cargoes and is potentially useful for developing a cell-permeable peptide library. Finally, we showed that, when a retro version of the carrier sequence was used, the peptide lost its translocating ability despite retaining a high content of a-helical structure in mimetic mem brane environments. This suggests that the propensity of peptides to a dopt a helical conformation is required but not sufficient for cellula r import and that other structural factors such as the side-chain topo logy of the carrier sequence are also important. Our studies together contribute to the more rational design of useful cell-permeable peptid es. (C) Munksgaard 1998.