As a remarkable pioneer in the field of studying protein-protein interactions (PPIs), Profacgen provides a novel technique, proximity-dependent biotin identification (BioID), which has been used for identification of proteins in the close vicinity of a protein of interest in living cells.
The introduction of BioID
Determining protein partners is an essential step toward understanding protein function and identifying relevant biological pathways. However, many conventional methods for investigating protein-protein interactions can be merely used in vitro, which may not reflect the actual interaction in native environments, or often encounter the loss of candidate proteins because of transient or weak protein interactions or protein insolubility.
BioID has emerged as a new tool based on enzyme-catalyzed proximity labeling (PL), to provide the possibility to study the spatial and interaction characteristics of proteins in living cells (Roux et al., 2012). Its main principle is that utilize the biotinylate proteins in proximity to a bait of interest that is fused to BirA. Once biotinylation, these proteins can be extracted from cell lysates, captured by streptavidin affinity purification, and identified by mass spectrometry. To date, BioID has been successfully applied to study all kinds of proteins and processes in cultured mammalian cells, mouse, plant, protoplasts, parasite, slime mold and yeast. It provides the possibility to better understand the complex function of individual proteins in vivo.
Figure 1. Schematic Diagram of BioID Technique
A Workflow of BioID
There are two main steps in a BioID process: (i) design and creation of a BioID fusion protein and validation of protein expression; (ii) biotinylation and identification of bait-interacting protein by mass spectrometry. The overview of the BioID procedure in detail as follows:
Figure 2. Diagram of Basic Protocol (Curr Protoc Cell Biol. 2016)
The Merits of BioID
Compared with conventional methods for the study of PPIs, BioID has become increasingly more favored by the researchers because of its own a variety of advantages, including:
In addition, it is worth noting that proximity biotinylation labeling is not as an evidence for direct and physical PPIs in BioID. Thus, Profacgen could also provide complementary testing services, like personalized IP assays, to further validate proximity interactions derived from BioID and determine which reflect the actual interaction in the physiological condition of cells.