Optogenetic tools that can finely control the space and time of protein-protein interaction are completely changing the experimental methods of cell biology. Most current optogenetic tools are created by using naturally occurring light dependent protein-protein interactions and fusing interacting partners into proteins of interest. However, since many photoactive proteins do not have known binding partners, the advantages of photogenetic control - accurate spatial and temporal resolution - are currently limited to some well-defined natural systems. In addition, the affinity and dynamics of natural interactions are usually not optimal and difficult to design in the absence of any structural information.
Creative BioMart provides a phage display strategy that can be used to find new binding partners that set the light and dark states of light switching proteins. These novel light-regulated protein-protein interactions can play a role in living cells to control the subcellular localization process.
A set of custom phages was constructed
Library design and screening of optical switch domain protein conjugates
We chose GA domain scaffold to make combinatorial phage display library. The scaffold Library (GA domain) was initially shown to fuse with the M13 coat protein pVIII, using a custom type 8 + 8 phage. Since this protein has multiple copies in each phage, characterization of light dependent protein-protein interaction with purified protein.
The target sequence was cloned into a customized expression vector.
A single phage clone was isolated from the third round phage library, and its binding specificity with different states of photoswitchable proteins and neutral avidin control wells was tested by standard ELISA protocol.
Expression and purification of protein binding agent
Finally, it was examined whether the found binding partners showed sufficient photoswitching affinity and selectivity to function in living cells.
We have extensive experience in handling dual hybrid systems. At the same time, there are a large number of prefabricated cDNA libraries for you to choose from.
We optimize the classic dual hybrid protocol and provide customers with specific customized solutions according to their different purposes.
We have more than 10 years of working experience in two hybrid research and experimental operation skills.
Creative BioMart is always devote us to provide high-quality and satisfactory service to our customers, if you are interested in our services or have some question, please feel free to contact us or make online inquiry.
