Intracellular delivery of small molecules, proteins, and nucleic acids
An amino acid sequence Arg-Lys-Met-Leu-Lys-Ser-Thr-Arg-Arg-Gln-Arg-Arg (SEQ ID NO:1) functions as a protein transduction domain (PTD) and is capable of delivering small molecules, proteins, and nucleic acids to an intracellular compartment of a cell. An amino terminal lysine linker improves the efficiency of the PTD. A nuclear localization signal can be used to target the PTD to a cell's nucleus. The PTD can be used in PTD-cargo moiety complexes that can reversibly immortalize cells and increase cell viability in culture.
1. An isolated and purified polypeptide with a protein transduction domain (PTD) which comprises Arg-Lys-Met-Leu-Lys-Ser-Thr-Arg-Arg-Gln-Arg-Arg (SEQ ID NO:1), wherein the polypeptide can traverse a cellular membrane.
2. The polypeptide of claim 1 further comprising at its amino terminus Lys-Xaa-Xaa, wherein Xaa is a small neutral polar or nonpolar amino acid.
3. The polypeptide of claim 2 comprising the amino acid sequence Lys-Gly-Gly-Arg-Lys-Met-Leu-Lys-Ser-Thr-Arg-Arg-Gln-Arg-Arg (SEQ ID NO:2).
4. The polypeptide of claim 1 further comprising a chemical cross-linker.
5. The polypeptide of claim 4 wherein the chemical cross-linker is maleimide or 3-nitro-2-pyridyldithio group.
6. A complex comprising a cargo moiety complexed with a polypeptide comprising a protein transduction domain (PTD) wherein said polypeptide is capable of complexing with said cargo moiety and wherein the PTD comprises Arg-Lys-Met-Leu-Lys-Ser-Thr-Arg-Arg-Gln-Arg-Arg (SEQ ID NO:1), and wherein the complex can traverse a cellular membrane.
7. The complex of claim 6 wherein the cargo moiety is selected from the group consisting of a small molecule, a nucleic acid, and a polypeptide.
8. The complex of claim 7 wherein the cargo moiety is a small molecule, wherein the small molecule is selected from the group consisting of a fluorescent marker, a dye, and a pharmaceutical agent.
9. The complex of claim 6 wherein said polypeptide comprising said PTD is a fusion protein.
10. The complex of claim 7 wherein the cargo moiety is a small molecule, wherein the small molecule comprises a radionuclide.
11. The complex of claim 6 wherein the cargo moiety is β-galactosidase or alkaline phosphatase.
12. A conjugate comprising a polypeptide with a protein transduction domain (PTD) linked to a cargo moiety, wherein the PTD comprises Arg-Lys-Met-Leu-Lys-Ser-Thr-Arg-Arg-Gln-Arg-Arg (SEQ ID NO:1), and wherein the conjugate can traverse a cellular membrane.
13. The conjugate of claim 12 , wherein the cargo moiety is selected from the group consisting of a small molecule, a nucleic acid, and a polypeptide.
14. The conjugate of claim 12 , wherein the conjugate comprises a fusion protein.
15. The conjugate of claim 12 , wherein the cargo moiety is β-galactosidase or alkaline phosphatase.