Orphan Receptor Strategy

As the number of identified natural ligands was rapidly outnumbered by the GPCRs discovered by DNA cloning, the orphan receptor strategy emerged in the mid-1990s [11]. This strategy, also referred to as "tissue-extract based approach," exposes orphan 7TM proteins to tissue extracts instead of purified ligands. Receptor activation is measured by changes in second messenger responses. Positive extracts are fractionated until the active component is isolated and characterized (Fig. 7.1). Despite low assay specificity, due to the complex composition of tissue extracts, the orphan receptor strategy proved to be useful for the identification of several natural ligands for orphan 7TM proteins. The first success of this strategy was the discovery of a novel transmitter for the ORL-1 receptor, which was cloned by homology to opioid receptors [20] . Based on the expression of ORL-1 in the central nervous system, brain tissue extracts were prepared and tested for their capacity to inhibit adenylyl cyclase activity in cells stably expressing ORL-1 receptors. A 17-residue long peptide sharing similarity with opioid peptides was ultimately isolated and named orphanin FQ or nociceptin (OFQ/N) 1 12, 21] . Subsequently, orphan 7TM proteins were screened randomly against large libraries of ligands [22] . These libraries typically contain all the ligands that have not been matched to any other receptor. Within a few years, more than 40 orphan 7TM proteins were deorphanized with this approach [23].

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