Chimeric proteins for use in transport of a selected substance into cells
First Claim
1. A chimeric protein comprising a functional domain and a carrier domain, wherein the functional domain comprises an amino acid sequence which binds a selected substance and the carrier domain comprises an amino acid sequence which binds a mammalian cell surface receptor other than the mammalian cell surface receptor which normally binds the selected substance in the mammal.
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Abstract
Chimeric proteins useful in transporting a selected substance present in extracellular fluids, such as blood or lymph, into cells; quantitative assays for the selected substance using chimeric proteins; DNA encoding the chimeric proteins; plasmids which contain DNA encoding the chimeric proteins; mammalian cells, modified to contain DNA encoding the chimeric proteins, which express and, optionally, secrete the chimeric proteins; a method of producing the chimeric proteins; a method of isolating the chimeric proteins; a method of using the chimeric proteins to assay the selected substance; and a method of reducing extracellular levels of the selected substance through administration of the chimeric protein, which results in transport of the selected substance into cells.
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Citations
42 Claims
- 1. A chimeric protein comprising a functional domain and a carrier domain, wherein the functional domain comprises an amino acid sequence which binds a selected substance and the carrier domain comprises an amino acid sequence which binds a mammalian cell surface receptor other than the mammalian cell surface receptor which normally binds the selected substance in the mammal.
- 7. DNA encoding a chimeric protein, the chimeric protein comprising a functional domain and a carrier domain, wherein the functional domain comprises an amino acid sequence which binds a selected substance and the carrier domain comprises an amino acid sequence which binds a mammalian cell surface receptor other than the mammalian cell surface receptor which normally binds the selected substance in the-mammal.
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10. DNA encoding a chimeric protein comprising a functional domain and a carrier domain, wherein the functional domain comprises the amino acid sequence of the ligand-binding domain of the human low density lipoprotein receptor and the carrier domain comprises an amino acid sequence which binds the human transferrin receptor.
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11. An expression plasmid comprising:
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a) a promoter;
b) DNA encoding a chimeric protein comprising a functional domain and a carrier domain, wherein the functional domain comprises an amino acid sequence which binds a selected substance and the carrier domain comprises an amino acid sequence which binds a mammalian cell surface receptor other than the mammalian cell surface receptor which normally binds the selected substance in the mammal, wherein expression of the DNA encoding the chimeric protein is under the control of the promoter.
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- 12. A genetically modified mammalian cell which expresses DNA encoding a chimeric protein, wherein the chimeric protein comprises a functional domain and a carrier domain, wherein the functional domain comprises an amino acid sequence which binds a selected substance and the carrier domain comprises an amino acid sequence which binds a mammalian cell surface receptor other than the mammalian cell surface receptor which normally binds the selected substance in the mammal.
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28. A method of transporting low density lipoprotein into a cell, comprising combining low density lipoprotein with:
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a) a chimeric protein which comprises a functional, domain and a carrier domain, wherein the functional domain comprises the amino acid sequence of the ligand-binding domain of human low density lipoprotein receptor and the carrier domain comprises an amino acid sequence which binds a human cell surface receptor other than human low density lipoprotein receptor; and
b) a human cell bearing a surface receptor which binds the amino acid sequence of the carrier domain of the chimeric protein, under conditions appropriate for binding of low density lipoprotein to the ligand binding domain of low density lipoprotein and for binding of the amino acid sequence of the carrier domain of the chimeric protein with the cell surface receptor on the human cell, whereby a complex of the chimeric protein and low density lipoprotein is formed and the complex is transported into the cell by endocytosis. - View Dependent Claims (29, 30)
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- 31. A method of lowering the level of low density lipoprotein in the bloodstream of an individual, comprising administering to the individual a chimeric protein comprising a functional domain and a carrier domain, wherein the functional domain comprises the amino acid sequence of the ligand-binding domain of human low density lipoprotein receptor and the carrier domain comprises an amino acid sequence which binds a human cell surface receptor other than human low density lipoprotein receptor, wherein low density lipoprotein in the bloodstream of the individual binds the ligand binding domain of low density lipoprotein in the chimeric protein and the carrier domain of the chimeric protein binds a human cell surface receptor, whereby the chimeric protein having low density lipoprotein bound thereto is transported into the cell bearing the cell surface receptor, thereby lowering the level of low density lipoprotein in the bloodstream of the individual.
- 36. A method of introducing a selected substance into a mammalian cell, comprising contacting a mammalian cell to a chimeric protein which binds the selected substance at a functional domain, and which binds a cell surface receptor on the mammalian cell at a carrier domain, such that the complex of the cell surface receptor, the chimeric protein, and the selected substance forms and the complex is bound to the cell surface receptor and transported into the cell by receptor-mediated endocytosis.
- 38. A method of introducing a functional protein domain into a mammalian cell, comprising contacting a mammalian cell with a chimeric protein which has a functional domain and which binds a cell surface receptor on the mammalian cell at a carrier domain, such that the complex of the cell surface receptor, the chimeric protein, and the selected substance forms and the complex is bound to the cell surface receptor and transported into the cell by receptor-mediated endocytosis.
Specification