Biomolecular optical sensors

• US 5,756,351 A
• Filed: 01/13/1997
• Issued: 05/26/1998
• Est. Priority Date: 01/13/1997
• Status: Expired due to Term

• Current Assignee: Regents of the University of California (University of California)
• Original Assignee: Regents of the University of California (University of California)
• Inventors: Mannuzzu, Lidia M., Moronne, Mario M., Isacoff, Ehud Y.
• Primary Examiner(s): Wax, Robert A.
• Assistant Examiner(s): Stole, Einar
• Application Number:
  US08/783,377
• Time in Patent Office: 498 Days
• Field of Search: 435/325, 435/252.3, 435/242, 435/254.11, 435/257.2, 435/410
• US Class Current: 435/325
• CPC Class Codes: C07K 14/705 Receptors; Cell surface antigens; Cell surface determinants tumour specific antigens C07K14/4748
• CPC Class Codes: G01N 33/54373 involving physiochemical end-point determination, e.g. wave-guides, FETS, gratings

Abstract

The invention provides protein-based sensors which report changes at a cell surface membrane. These sensors comprise an endogenous cell surface protein having a post-translationally generated luminescer at a predetermined residue. In operation, the protein adopts one of a plurality of different interconvertable signal-dependent conformations, whereunder the luminescer provides corresponding different luminescence. The cells comprise transgenes encoding the protein sensor which is translated from such transgenes by the host cell'"'"'s machinery with natural amino acids and expressed on the cell surface. After surface expression, the cell is contacted with a luminescer generating reagent which generates a luminescer at a predetermined residue of the protein. A wide variety of luminescers and chemistries for generating the selected luminescer at the target residue of the sensor protein may be used.

Claims

1. A cell comprising an artificial molecular optical sensor, said sensor comprising a predetermined transmembrane cell surface protein comprising a post-translationally generated luminescer at a predetermined residue of said protein, wherein said protein adopts one of a plurality of different interconvertable signal-dependent conformations, whereunder said luminescer provides corresponding different luminescence, wherein said cell comprises a transgene encoding said protein and said cell expresses said protein said residue is located within about 10 residues of a transmembrane domain--extracellular domain junction of said protein.

2. A cell according to claim 1 wherein said protein is a membrane channel protein, a receptor or a signal transducing protein.

3. A cell according to claim 1 wherein said signal is the binding of a ligand to said protein or a change in a charge distribution across the plasma membrane of said cell at said protein.

4. A cell according to claim 1 wherein said residue is not native to said protein.

5. A cell according to claim 1 wherein said residue comprises a functionally available sulfhydryl group.

6. A cell according to claim 1 wherein said residue comprises a functionally available sulfhydryl group and other otherwise functionally available sulfhydryl groups of said cell surface are chemically blocked.

7. A cell according to claim 1 wherein said cell comprises a blocking reagent at residues which would otherwise comprise said luminescer.

8. A cell according to claim 1 wherein said protein is a membrane ion channel protein.