Recombinant renilla reniformis system for bioluminescence resonance energy transfer

A composition comprising a first recombinant fusion protein comprising a first polypeptide fused to the amino acid sequence of R. reniformis luciferase and a second recombinant fusion protein comprising a second polypeptide fused to the amino acid sequence of R. reniformis GFP.

A method of detecting protein:

• protein interactions, said method comprising:
• a) contacting a first fusion protein comprising a first polypeptide fused to R. reniformis luciferase polypeptide, and a second fusion protein comprising a second polypeptide fused to R. reniformis GFP polypeptide and a substrate for luciferase under conditions sufficient to permit BRET;
• b) detecting BRET induced fluorescent emission from said R. reniformis GFP, wherein said fluorescent emission from said R. reniformis GFP indicates protein:protein interaction between said first and said second polypeptides.

A method of determining the location in a cell of a protein:protein interaction between two polypeptides, said method comprising the steps of:

• a) introducing into a cell a first polynucleotide sequence encoding a first polynucleotide comprising an expression cassette containing a sequence encoding R. reniformis luciferase and a second polynucleotide comprising an expression cassette containing a sequence encoding R. reniformis GFP;
• b) contacting said cell of step a) with a substrate for luciferase;
• c) detecting BRET induced fluorescent emission from said R. reniformis GFP as an indicator of the location of the cellular compartment where said protein:protein interaction between said first and said second polypeptides occurs.

A method of identifying cells in which there is a protein:protein interaction between two polypeptides of interest, said method comprising the steps of:

• a) introducing into a population of cells a first polynucleotide sequence encoding a first polynucleotide comprising an expression cassette containing a sequence encoding R. reniformis luciferase and a second polynucleotide comprising an expression cassette containing a sequence encoding R. reniformis GFP;
• b) contacting said cell of step a) with a substrate for luciferase;
• c) detecting BRET induced fluorescent emission from said R. reniformis GFP, wherein said fluorescent emission from said R. reniformis GFP identifies a cell in which a protein:protein interaction between said first and said second polypeptides occurs.

A method of screening for a candidate modulator that modulates a protein:protein interaction between two polypeptides, said method comprising:

• a) contacting a first fusion protein comprising a first polypeptide and R. reniformis GFP polypeptide, and a second fusion protein comprising a second polypeptide and R. reniformis luciferase polypeptide, a candidate modulator, under conditions that permit binding of said fusion polypeptides to each other, and a substrate for luciferase;
• b) measuring BRET induced fluorescent emission, wherein said fluorescent emission indicates a protein:protein interaction between said first and said second polypeptides and;
• c) comparing the amount of fluorescence emission in step b) to the amount of fluorescence emission in the absence of said candidate modulator.

A kit for detecting a protein:protein interaction, said kit comprising a first polynucleotide comprising an expression cassette containing a sequence encoding R. reniformis luciferase and a second polynucleotide comprising an expression cassette containing a sequence encoding R. reniformis GFP and packaging materials therefore wherein the first and second polynucleotides are packaged independently.

A kit for comprising a polynucleotide encoding a R. reniformis luciferase polypeptide and a R. reniformis GFP polypeptide, and packaging materials therefore.