Design of novel chimeric GPCRs: Identification of equivalent regions at the sequence, structural and biophysical level

Currently, 60% of the druggable G-Protein Coupled Receptors (GPCRs), proteins involved in a broad range of diseases, remain untargeted due to the lack of insights at their sequence and structural level. Chimeric GPCRs, typically the merger of well- and not well-characterized GPCRs, can help to increase GPCR understanding by revealing novel 3D structures, biological function, and ligand binders. However, designing chimeras is challenging as equivalent regions need to be merged to ensure expression and function of the resulting chimera.
Correct sequence alignment of the GPCRs is thus crucial. Therefore, we evaluated the state-of-the-art Multiple Sequence Alignment (MSA) of GPCRs by studying the predicted biophysical properties of 312 aligned class A GPCR sequences. Unexpected shifts in predicted biophysical behaviors were observed for GPCRs known to be phylogenetically related.
Based on an AlphaFold2-guided structure-based MSA, an improved MSA was obtained where specific residues known to be equivalent were aligned, and where biophysical behaviors of related GPCRs were better matched. This work will lead to proposing novel chimeric designs and exploring a method to improve sequence alignment directly using the predicted biophysical behavior.