Galphaq protein variants and their use in the analysis and discovery of agonists and antagonists of chemosensory receptors

Abstract

The invention provides a series of Gα q protein variants that functionally couple to sensory cell receptors such as taste GPCRs (TRs) and olfactory GPCRs (ORs) in an overly promiscuous manner. According to the invention, the functional coupling can be determined, for example, by measuring changes in intracellular IP3, or calcium. In a particular embodiment, the Gα q protein variants can be expressed in mammalian cell lines or Xenopus oocytes, and then evaluated using calcium fluorescence imaging and electrophysiological recording.

1 - 39 . (canceled)  
     
     
         40 . An isolated nucleic acid sequence encoding a G q  protein variant wherein at least the last five amino acids are replaced by the 5 C-terminal amino acids of Gα olf  or transducin and the resultant altered G q  variant exhibits greater promiscuity compared to the corresponding native G q  protein.  
     
     
         41 . An isolated nucleic acid sequence encoding the G q  protein variant of  claim 40 , wherein the native G q  protein is a human G q  protein.  
     
     
         42 . An isolated nucleic acid sequence encoding the G q  protein variant of  claim 40 , wherein the native G q  protein is a mouse G q  protein.  
     
     
         43 . An isolated nucleic acid sequence encoding a G q  protein variant comprising a nucleic acid encoding a polypeptide with at least 95% amino acid sequence identity to a sequence selected from the group consisting of SEQ ID NOS 1-26.  
     
     
         44 . An isolated nucleic acid sequence encoding a Gα q  protein variant comprising a nucleic acid encoding polypeptide with greater than 95% amino acid sequence identity to a sequence selected from the group consisting of SEQ ID Nos 1-26.  
     
     
         45 . An isolated nucleic acid sequence encoding a Gα q  protein variant that is encased by a amino sequence selected from the group consisting of SEQ ID Nos 1-26.  
     
     
         46 . An expression vector comprising the nucleic acid sequence of  claim 40  operably linked to a promoter that functions in mammalian cells or  Xenopus  oocytes.  
     
     
         47 . An expression vector comprising the nucleic acid sequence of  claim 41  operably linked to a promoter that functions in mammalian cells or  Xenopus  oocytes.  
     
     
         48 . An expression vector comprising the nucleic acid sequence of  claim 42  operably linked to a promoter that functions in mammalian cells or  Xenopus  oocytes.  
     
     
         49 . An expression vector comprising the nucleic acid sequence of  claim 43  operably linked to a promoter that functions in mammalian cells or  Xenopus  oocytes.  
     
     
         50 . An expression vector comprising the nucleic acid sequence of  claim 44  operably linked to a promoter that functions in mammalian cells or  Xenopus  oocytes.  
     
     
         51 . An expression vector comprising the nucleic acid sequence of  claim 45  operably linked to a promoter that functions in mammalian cells or  Xenopus  oocytes.  
     
     
         52 . A host cell comprising the expression vector of  claim 46 .  
     
     
         53 . A host cell comprising the expression vector of  claim 47 .  
     
     
         54 . A host cell comprising the expression vector of  claim 48 .  
     
     
         55 . A host cell comprising the expression vector of  claim 49 .  
     
     
         56 . A host cell comprising the expression vector of  claim 50 .  
     
     
         57 . A host cell comprising the expression vector of  claim 51.