Patent ID: 10456475

Abstract:
Cell penetrating peptides (CPPs) are established as a strategy to move cargoes into the interior of eukaryotic cells by engaging import machinery on the cell surface. In most cases the CPP is covalently linked to the cargo; it is common to express cargo proteins with a CPP extension. In some experiments a non-specific interactions (e.g., hydrophobic interactions) have been used to form CPP-cargo complexes, but this has numerous drawbacks. Transport is less efficient and the lack of specificity means that other macromolecules in the medium will also be internalized. This application describes the use of specific CPP labeled adaptor proteins that can be used to move a wide variety of cargoes into the cell interior. The prototype adaptor protein is calmodulin; it is small, stable and easily produced, and binds short (17 amino acid) targets with high affinity in the presence of calcium. A cargo produced with a calmodulin binding tag can be internalized efficiently by CPP tagged calmodulin. Because calcium in the cell interior is typically kept low, CPP-calmodulin releases cargoes inside the cell. The calmodulin binding tag can be used to efficiently purify target proteins by affinity chromatography.