A Cellular Assay for the Identification and Characterization of Connexin Gap Junction Modulators.

Connexin gap junctions (Cx GJs) enable the passage of small molecules and ions between cells and are therefore important for cell-to-cell communication. Their dysfunction is associated with diseases, and small molecules acting as modulators of GJs may therefore be useful as therapeutic drugs. To identify GJ modulators, suitable assays are needed that allow compound screening.

Adenosine A receptor ligand recognition and signaling is blocked by A receptors.

The adenosine receptor (AR) subtypes A and A are rhodopsin-like G protein-coupled receptors whose expression is highly regulated under pathological, e.g. hypoxic, ischemic and inflammatory conditions.

Quantification of green fluorescent protein-(GFP-) tagged membrane proteins by capillary gel electrophoresis.

A fast and robust procedure for the quantification of GFP-tagged membrane proteins in cell homogenates was developed employing capillary gel electrophoresis coupled to laser-induced fluorescence detection (CGE-LIF). The new method was found to be highly sensitive and applicable to structurally diverse membrane proteins including synaptic vesicle protein 2A (SV2A), adenosine A receptor (AAR), and connexin 43 (Cx43). Quantification of SV2A and AAR using radioligand binding assays confirmed the results obtained with CGE-LIF.

Interaction of Approved Drugs with Synaptic Vesicle Protein 2A.

Levetiracetam (LEV) and its recently approved derivative brivaracetam are anti-epileptic drugs with a unique mechanism of action. The synaptic vesicle protein 2A (SV2A) was previously identified as their main target. In the current study, we tested a collection of 500 approved drugs for interaction with the human SV2A protein expressed in Chinese hamster ovary cells.