Two-hybrid assays refer to a set of molecular techniques used in genetics and molecular biology research to investigate protein-protein interactions in living cells. The term "two-hybrid" indicates the two distinct components of the assay that allow the detection of such interactions.
The first component is the "bait" protein, which is typically a protein of interest that is tagged with a DNA-binding domain. This bait protein is typically fused to a DNA-binding domain derived from a transcription factor, which allows it to bind to a specific DNA sequence element.
The second component is the "prey" protein, which is another protein of interest, or a library of proteins, that is fused to a transcriptional activation domain. This prey protein library can potentially contain all the proteins present in a given cell or organism.
When the bait and prey proteins interact in a living cell, they bring the DNA-binding and transcriptional activation domains together, leading to the transcription of a reporter gene. This reporter gene encodes a protein that can be easily detected or quantified, such as an enzyme or a fluorescent protein.
By identifying the cells or organisms in which the reporter gene is activated, researchers can infer protein-protein interactions. Two-hybrid assays are commonly used to probe protein interaction networks, identify novel protein partners, and investigate the effects of mutations on protein interactions.
Overall, two-hybrid assays provide a powerful tool for studying protein-protein interactions, allowing researchers to gain insights into the complex molecular machinery underlying cellular processes.
