In-depth identification of protein interaction partners is an important method for studying protein functions and positioning them in the intracellular interaction network. Over the years, various biotechnologies have been developed to meet this goal, including some methods specifically for studying post-translational modifications of mammalian proteins. Although these methods can clarify their inherent physiological context, they also have some inherent limitations.
Nowadays, these technologies have undergone many changes, providing a more appropriate environment for studying the interactions between mammalian proteins. Mammalian Protein-Protein Interaction Trap (MAPPIT) is a new type of mammalian two-hybrid system protocol based on the insights of type I cytokine receptor signal transduction, that is, activated JAK kinase can phosphorylate STAT recruitment sites in trans. The bait protein is fused with the mutant receptor chimera, and STAT target site has been eliminated from it. The prey protein is linked to a receptor fragment containing a functional STAT recruitment site. When the bait-prey interaction, the prey chimeras are phosphorylated, and the recruited STAT molecule is activated and induces the transcription of the reporter gene under the control of the STAT-responsive promoter. Compared with other similar methods, MAPPIT provides the best physiological environment, and has the comprehensive advantages of separation of interaction zone and effect zone and rapid identification of false positives.
Figure 1. Schematic representation of the JAK-STAT pathway (Eyckerman, S.; et al.)
Creative Proteomics has recruited many experienced technicians to provide high-quality protein-protein interaction research services to researchers all over the world. The MAPPIT system we established has been successfully applied in many studies, providing a normal physiological environment for the mammalian protein to be tested. This system ensures the correct folding of the protein and provides various necessary cofactors and regulatory proteins participating in post-translational modifications. In order to ensure the accuracy of the experiment, our technicians can also offer experimental options such as the determination of the optimal selective puromycin concentration before screening, subcloning the prey into the vector for individual testing and confirming the interaction, and reducing the expression level of the prey to reduce non-specific binding.
Figure 2. Flow chart of the MAPPIT screening procedure (Eyckerman, S.; et al.)
Customers can choose different technology platforms according to project requirements, or contact us directly for consultation, and our expert team will provide you with customized experimental procedures.
Creative Proteomics is an international biotechnology company dedicated to research in molecular interactions and other related fields. The mammalian protein-protein interaction trap platform we constructed has the characteristics of high quality and efficiency, and the data obtained can be directly used for paper publication. Our one-stop service aims to save customers time and money.