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Mammalian Two-hybrid Service

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Creative Biolabs offers mammalian two-hybrid service, which allows rapid and convenient analysis of protein-protein interactions in transfected mammalian cells. Compared with the yeast two-hybrid system, the mammalian two-hybrid system provides the milieu for the bona fide posttranslational modification and localization of most eukaryotic proteins and, therefore, should be a better choice to study proteins of a mammalian origin.

Two-hybrid systems are extremely powerful methods for detecting protein-protein interactions in vivo. However, a major drawback of testing protein-protein interactions in a heterologous system such as the yeast is that interactions may depend on certain posttranslational modifications such as disulfide bridge formation, glycosylation, or phosphorylation, which may not occur properly or at all in the yeast system. In addition, since the fusion proteins in the two-hybrid system must be targeted to the nucleus, extracellular proteins or proteins with stronger targeting signals may be at a disadvantage. It has been shown that sub-domains of proteins may interact better than full length clones in yeast two-hybrid systems, perhaps due to the lack of certain folding restraints. What’s more, certain proteins may become toxic when expressed in the yeast two-hybrid system or that targeted to the nucleus may become toxic. Other proteins may degrade essential yeast proteins or proteins whose presence are required for the assay. Such genes may be counter-selected for during growth and may result in problems.

The mammalian two-hybrid system is a very powerful tool to investigate protein-protein interactions in terms of functional domains and identify potential binding ligands and partners of a protein. Creative Biolabs has adapted yeast two-hybrid system for use in mammalian cells. In our system, one protein (X) is fused to a DNA-binding domain interacts with a second protein (Y), which is fused to a transcriptional activation domain. A luciferase reporter gene was introduced as the readout of interaction between proteins X and Y.

One major advantage of our mammalian two-hybrid system over yeast systems is that mammalian protein interactions can be studied in an environment which is that is more similar to that in vivo. Our mammalian two-hybrid service can be applied in confirming suspected interactions between two proteins, identifying residues/domains involved in protein-protein interactions and identifying small molecules that affect protein-protein interactions, ect.




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