Commonwealth Scientific and Industrial Research Organisation

Virginia Leitch

Stephen Charles Trowell

Helen Dacres

Irene Horne

Mira Maria Dumancic

Irene Mary Horne

The present invention relates to methods and polypeptides for detecting a compound in a sample. In particular, the present invention relates to the use of a cell-free composition comprising at least one G protein coupled receptor embedded in a lipid Mayer which when expressed in a cell the N-terminus of the G protein coupled receptor, or subunits thereof, is outside the cell and the C-terminus is inside the cell, and which is capable of binding the compound. Optionally, the composition also comprises at least one accessory molecule that directly or indirectly binds an intracellular loop and/or the C-terminus of the G protein coupled receptor. The G protein coupled receptor, and/or accessory molecule when present, in combination comprise a bioluminescent protein and an acceptor molecule, which enables bioluminescent resonance energy transfer (BRET) to be used to detect the compound binding the receptor.

Measuring G protein coupled receptor activation

Nucleotide and amino acid sequences of luciferase peptides that are encoded by genes within the genome of(Diptera) are disclosed. Specifically provided are functional ATP-dependent luciferases that catalyze luminescence reactions with emission spectra within the blue portion of the spectrum. The present invention specifically provides isolated peptide and nucleic acid molecules, methods of identifying orthologs and active subsequences of the enzyme peptides, and methods of identifying modulators and substrates of the luciferase peptides. Methods of assays, including multiple reporter assays utilizing at least two ATP-dependent luciferases are provided.

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Belconnen, Australia

Virginia Leitch