Cell penetrating peptides facilitate protein transport into the cell-sized asymmetric lipid vesicles at arbitrary timing.

Abstract

Cell penetrating peptides (CPP) play an important role of drug delivery system. CPP have a strategy to overcame the membrane impermeability. There are two pathways to across the lipid bilayer via CPPs such as direct translocation and endocytosis. The direct translocation of CPPs was investigated using an artificial lipid membrane, such as planar lipid bilayers and nano-sized lipid vesicles. The direct translocation via CPPs using the planar lipid bilayers suggests that the negative transmembrane potential generating between planar lipid bilayers plays an important role for the direct translocation of β-gal via pep-1. In this study, the interactions between CPPs and cell-sized lipid vesicles were investigated using two peptides (Pep-1, penetratin). Next, the interaction between the CPP-DNase I complexes and membrane were observed. In addition, we investigated the DNase I translocation of CPP-DNase I complexes into the GUVs. Finally, we demonstrated the formation of a cross-linked actin network in GUVs using CPP-mediated translocation.CPPs-DNase I complexes was easily interacted with the asymmetric GUVs containing DOPG or DOPS in the inner leaflet. Therefore, CPPs mediated direct translocation of DNase I into the asymmetric GUVs with negatively charged lipids on the inner leaflet. These results indicate that the CPPs-mediated direct translocation could be triggered by the transmembrane potential generating by asymmetric GUV membrane. We found that the penetratin-mediated translocation of DNase I was more efficient than the Pep-1-mediated translocation. Using this asymmetric membrane component and CPP condition, we could translocate streptavidin into GUVs. Thus, highly versatile of CPP-mediated translocation was shown. This system will contribute to creations of well-defined artificial cell models with the capacity to control the start of inside reaction.