cAMP assay for GPCR ligand characterization: application of BacMam expression system.

Cyclic adenosine monophosphate (cAMP) is a second messenger of many G-protein-coupled receptors. The change in cellular cAMP level has widely been used to estimate the biological activity of various GPCR-specific agents. Förster resonance energy transfer (FRET) biosensors have been around for almost 10 years and became increasingly popular for cAMP detection.

1-substituted apomorphines as potent dopamine agonists.

A novel set of 1-substituted apomorphines as dopaminergic agonists were synthesized according to our new strategy employing the acid-catalyzed rearrangement of diversely functionalized 5β-substituted-6-demethoxythebaines. The activities of new compounds for dopamine receptors subtypes were evaluated using HEK293 based stable cell lines expressing D1, D2L or D3 receptor subtypes. All studied compounds had affinities in nanomolar range for D2L and D3 receptors and the change of the nature of substituent in position 1 had only moderate effect.

BacMam system for FRET-based cAMP sensor expression in studies of melanocortin MC1 receptor activation.

Cyclic adenosine monophosphate (cAMP) is a second messenger of many G-protein-coupled receptors (GPCRs) and a useful readout molecule to estimate the biological activity of various GPCR-specific agents. Here we report the development and use of a Förster resonance energy transfer (FRET) biosensor for cAMP (Epac2-camps) combined with a baculovirus-based BacMam transduction system. The constructed BacMam-Epac2-camps viral transduction system is a simple and robust tool for ligand screening at the second-messenger level in a variety of mammalian cell lines.