DiscoverX cell-based assays feature the powerful in vivo application of the established Enzyme Fragment Complementation technology. In this approach, two complementing fragments (ProLink, a small enzyme donor peptide fragment, and EA, a larger enzyme acceptor fragment) of the β-galactosidase (β-gal) enzyme are expressed as protein fusions in stably transfected, clonally derived cells.
The pCMV-ProLink™ cloning vectors are intended for cloning a GPCR or other protein of interest with ProLink as a C-terminal tag, and subsequently transfecting into a parental cell line containing an EA-tagged protein. ProLink 1 peptide tag has low affinity for EA and is useful for strong protein:protein interactions, while ProLink 2 peptide tag has a higher affinity (~3-fold greater) for EA and is useful for weak protein:protein interactions. ARMS (Arrestin Recruitment Modifying Sequence) ProLink tags contain GRK (G-protein Receptor Kinase) consensus phosphorylation sites and can improve assays that have a low signal-to-background ratio.