Faculty
Science
Supervisor Name
Peter Chidiac
Keywords
receptors, luciferin, allosteric site, pharmaceuticals, sodium pocket
Description
G protein-coupled receptors (GPCRs) are proteins that are important in physiological regulatory processes within the body, and for this reason are important drug targets. When bound to an agonist, such as neurotransmitters or hormones, the receptor adopts an active state to allow these biochemical pathways to occur. However, mutations can arise within the receptor that affect its ability to bind its agonist. This natural variation found within the genome can make it difficult to design pharmaceuticals to target the receptors.
To see the effects of these point substitutions on agonist-induced receptor activation, mutations were made within a negative allosteric site of the Adenosine Receptor A2A called the sodium ion binding pocket. It was hypothesized that these mutations destabilize the sodium ion, increasing the ability of an agonist to bind the Orthosteric site. These point substitutions were modelled using an online protein visualization program (PyMOL), and any shifts in residue position were noted.
An in vitro analysis was then conducted to fully understand this concept, by studying the effects of the transfection of natural human variants into cells. A GloSensor cAMP assay was used, which produced luminescence activity upon receptor activation. Compared to wild type proteins, receptor activity was found to have decreased with mutation S91A. These findings suggest that receptor activation is more complex than originally thought, and that more research must be conducted to better understand these effects on pharmaceuticals.
Acknowledgements
I would like to thank the Chidiac lab for their support throughout the summer. A special thank you to Fang (Mandy) Wang as well as Garmen Shelly Ng for their supervision and guidance.
Creative Commons License
This work is licensed under a Creative Commons Attribution-No Derivative Works 4.0 License.
Document Type
Poster
Included in
Medical Pharmacology Commons, Medical Physiology Commons, Pharmaceutics and Drug Design Commons
Point Substitutions in G Protein-Coupled Receptors
G protein-coupled receptors (GPCRs) are proteins that are important in physiological regulatory processes within the body, and for this reason are important drug targets. When bound to an agonist, such as neurotransmitters or hormones, the receptor adopts an active state to allow these biochemical pathways to occur. However, mutations can arise within the receptor that affect its ability to bind its agonist. This natural variation found within the genome can make it difficult to design pharmaceuticals to target the receptors.
To see the effects of these point substitutions on agonist-induced receptor activation, mutations were made within a negative allosteric site of the Adenosine Receptor A2A called the sodium ion binding pocket. It was hypothesized that these mutations destabilize the sodium ion, increasing the ability of an agonist to bind the Orthosteric site. These point substitutions were modelled using an online protein visualization program (PyMOL), and any shifts in residue position were noted.
An in vitro analysis was then conducted to fully understand this concept, by studying the effects of the transfection of natural human variants into cells. A GloSensor cAMP assay was used, which produced luminescence activity upon receptor activation. Compared to wild type proteins, receptor activity was found to have decreased with mutation S91A. These findings suggest that receptor activation is more complex than originally thought, and that more research must be conducted to better understand these effects on pharmaceuticals.