Patent ID: 6395478
Filing Date: 2002-05-28
Classification: B01J,B82Y,C12N,C40B,G01N,G16B

Abstract:
A method of detecting one or more protein-protein interactions comprising:(a) recombinantly expressing within a population of host cells a first population of first fusion proteins, each said first fusion protein comprising a first protein sequence and a DNA binding domain in which the DNA binding domain is the same in each said first fusion protein, in which said host cells comprise at least one polynucleotide operably linked to a promoter driven by one or more DNA binding sites recognized by said DNA binding domain of said first fusion protein such that an interaction of a first fusion protein with a second fusion protein, said second fusion protein comprising a transcriptional regulatory domain which is an activation domain, results in activation of transcription of said at least one polynucleotide by said activation domain, and in which said first population of first fusion proteins has a complexity of at least 10; (b) negatively selecting to eliminate those host cells expressing said first population of first fusion proteins in which said activation of transcription of said at least one polynucleotide occurs in the absence of a second fusion protein, said negatively selecting being performed by a method comprising incubating said host cells expressing said first population of first fusion proteins in an environment in which substantial death of said host cells occurs if said activation of transcription occurs; (c) after step (b), recombinantly co-expressing with said population of host cells a second population of second fusion proteins, each said second fusion protein comprising a second protein sequence and an activation domain of a transcriptional regulator, in which the activation domain is the same in each said second fusion protein, and in which said second population of second fusion proteins has a complexity of at least 10; and (d) detecting said activation of transcription of said at least one polynucleotide, thereby detecting cells in which an interaction between a first protein and a second protein has occurred, wherein the first and second populations of fusion proteins comprise first and second protein sequences, respectively, that are encoded by DNA sequences which are drawn from different DNA populations.