Structural determinants of dopamine receptor agonist selectivity

Dopamine receptors (DRs) are G protein-coupled receptors (GPCRs) expressed in the central nervous system. When activated, DRs can trigger different downstream signal transduction cascades based on which they are divided into two major classes: D1-like, which includes excitatory D1 and D5 receptors, and D2-like, including inhibitory D2, D3 and D4 receptors. Drugs acting on DRs are used to treat several neurological disorders, like Parkinson´s disease (PD), schizophrenia, depression or bipolar disorders. PD, for example, is caused by the early death of dopaminergic neurons in substantia nigra pars compacta. Therapeutic options for PD acting on DRs include the D2-like selective agonists ropinirole and pramipexole. Despite their approval over two decades ago, a comprehensive investigation of the structural determinants responsible for their D2-like DR selectivity has not yet been conducted. In this work we therefore aim to elucidate the molecular determinants conferring (un)selective binding of DR agonists, including dopamine, pramipexole and ropinirole. For this purpose, we investigated DRs signaling cascades through 16 Galpha-proteins (collectively named the transducerome) using bioluminescence resonance energy transfer (BRET) biosensors and a luminescence based cAMP assay. Computational analyses allowed us to perform information driven site-directed mutagenesis. Selected mutations in D2 and D1 receptors modified the efficacy and/or potency of pramipexole and dopamine dependent activation in comparison to wild-type receptors. In this study, we were able to identify hotspot mutations within the binding pocket of D1 and D2, which are involved in D2-like pramipexole selectivity, efficacy and potency.