Patent ID: 8076159

Claim:
A method of detecting a ligand in vitro, comprising: providing a ligand-regulable transactivation system comprising: a reporter polynucleotide that comprises a binding sequence, a promoter sequence, and a reporter sequence encoding a detectable bioluminescent protein, wherein the binding sequence is connected with the promoter sequence and the promoter sequence is connected with the reporter sequence; and an activator fusion protein that comprises: a DNA binding domain capable of interacting with the binding sequence of the reporter polynucleotide, an estrogen receptor folding domain (ER folding domain) capable of binding a ligand, wherein the ER folding domain is selected from: a sequence consisting of SEQ ID NO:1 (human estrogen receptor, alpha, amino acids 281-549), a sequence consisting of SEQ ID NO:2 (human estrogen receptor, alpha, amino acids 281-595), a sequence consisting of SEQ ID NO:4 (mouse estrogen receptor, alpha, amino acids 281-549), a sequence consisting of SEQ ID NO:5 (mouse estrogen receptor, alpha, amino acids 281-599), SEQ ID NO:23 and SEQ ID NO:24, and a transactivation domain capable of interacting with the promoter sequence of the reporter polynucleotide, wherein the DNA binding domain is connected to the estrogen receptor folding domain, and the estrogen receptor folding domain is connected with the transactivation domain, and wherein interaction of the DNA binding domain and the transactivation domain of the activator fusion protein with the binding sequence and promoter sequence of the reporter polynucleotide regulates expression of the detectable bioluminescent protein; introducing a ligand to the system; and detecting a bioluminescent signal in the presence of a bioluminescence initiating compound if the ligand causes the ER folding domain to change from a non-interacting conformational position to an interacting conformational position, wherein the interacting conformational position positions the DNA binding domain and the transactivation domain of the activator fusion protein so that both interact with the binding sequence and the promoter sequence of the reporter polynucleotide, which causes expression of the detectable bioluminescent protein encoded by the reporter sequence.