The coiled-coil protein carrier structure affects the activation of certain endocytosis pathways.

Coiled-coil protein carrier (CCPC) 140 is a rigid and anisotropically structured cationic coiled-coil artificial protein that has displayed up to a 1000 times higher level of cellular internalization activity than that of unstructured cell-penetrating peptides. Previous studies have demonstrated that CCPC 140's rigid and anisotropic structural properties and cationic surface properties are important for its superior cellular internalization activity. In this study, we investigated whether each physicochemical characteristic of CCPC 140 effectively contributed to activating the cellular internalization pathway. By evaluating CCPC 140's ability to penetrate glycosaminoglycan (GAG)-lacking cells, the activation of GAG-dependent endocytosis by electrostatic interactions between cationic CCPC 140 and anionic GAGs has been found to play a major role in CCPC 140's superior cellular internalization activity. Using endocytosis inhibitors, it was revealed that the GAG-binding-dependent activation of caveola-mediated endocytosis plays a role in cellular internalization, which requires rigid and anisotropic structural properties, not the cationic properties of CCPC 140. Macropinocytosis is a common route of cellular internalization. However, CCPC 140's rigid and anisotropic structural properties activate macropinocytosis, but this does not involve the Rho-family GTPase-dependent macropinocytosis pathway.