Interaction trap systems for detecting protein interactions

A method of detecting an interacting protein in a population of proteins, comprising:

• (a) providing a host cell which contains
• (i) a reporter gene operably linked to a DNA-binding-protein recognition site;
• and
• (ii) a fusion gene which expresses a fusion protein, said fusion protein comprising a test protein covalently bonded to a binding moiety which is capable of specifically binding to said DNA-binding-protein recognition site;
• (b) introducing into said host cell a second fusion gene which expresses a second fusion protein, said second fusion protein comprising one of said population of proteins covalently bonded to a gene activating moiety and being conformationally-constrained;
• and
• (c) measuring expression of said reporter gene.

A method of identifying a candidate interactor, comprising:

• (a) providing a reporter gene operably linked to a DNA-binding-protein recognition site;
• (b) providing a first fusion protein, said first fusion protein comprising a first protein covalently bonded to a binding moiety which is capable of specifically binding to said DNA-binding-protein recognition site;
• (c) providing a second fusion protein, said second fusion protein comprising a second protein covalently bonded to a gene activating moiety and being conformationally-constrained, said second protein being capable of interacting with said first protein;
• (d) contacting said candidate interactor with said first protein and/or said second protein;
• and
• (e) measuring expression of said reporter gene.

A method of assaying an interaction between a first protein and a second protein, comprising:

• (a) providing a reporter gene operably linked to a DNA-binding-protein recognition site;
• (b) providing a first fusion protein comprising said first protein covalently bonded to a binding moiety which is capable of specifically binding to said DNA-binding-protein recognition site;
• (c) providing a second fusion protein comprising said second protein covalently bonded to a gene activating moiety and being conformationally-constrained;
• (d) combining said reporter gene, said first fusion protein, and said second fusion protein;
• and
• (e) measuring expression of said reporter gene.

A protein isolated by a method comprising:

• (a) providing a host cell which contains
• (i) a reporter gene operably linked to a DNA-binding-protein recognition site;
• and
• (ii) a fusion gene which expresses a fusion protein, said fusion protein comprising a test protein covalently bonded to a binding moiety which is capable of specifically binding to said DNA-binding-protein recognition site;
• (b) introducing into said host cell a second fusion gene which expresses a second fusion protein, said second fusion protein comprising one of said population of proteins covalently bonded to a gene activating moiety and being conformationally-constrained;
• and
• (c) measuring expression of said reporter gene;
• and
• (d) isolating a protein based on its ability to alter the expression of said reporter gene when present in said second fusion protein.

An interactor protein isolated by a method comprising:

• (a) providing a reporter gene operably linked to a DNA-binding-protein recognition site;
• (b) providing a first fusion protein, said first fusion protein comprising a first protein covalently bonded to a binding moiety which is capable of specifically binding to said DNA-binding-protein recognition site;
• (c) providing a second fusion protein, said second fusion protein comprising a second protein covalently bonded to a gene activating moiety and being conformationally-constrained, said second protein being capable of interacting with said first protein;
• (d) contacting a candidate interactor protein with said first protein or said second protein;
• (e) measuring expression of said reporter gene;
• and
• (f) isolating an interactor protein based on its ability to alter the expression of said reporter gene when present with said first protein or said second protein.

A method of assaying an interaction between a first protein and a second protein, comprising:

• (a) providing said first protein;
• (b) providing a fusion protein comprising said second protein, said second protein being conformationally-constrained;
• (c) contacting said first protein with said fusion protein under conditions which allow complex formation;
• (d) detecting said complex as an indication of an interaction;
• and
• (e) determining whether said first protein interacts with said fusion protein inside a cell.