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Kinetic determinants of GPCR signaling: from ultra-fast to diffusion-limited

Stanford Neurosciences Institute, Rabindra Shivnaraine

G protein-coupled receptors (GPCRs) are proteins that exist within the cell membrane and act to transfer the information encoded within neurotransmitters and drugs into cell responses. GPCRs exist throughout the body in several systems including the nervous system.  Drugs targeting GPCRs account for approximately 40% of all current pharmaceuticals.  These drugs cause GPCRs to undergo structural changes known as conformational changes which are not static but dynamic.  That is, they are constantly changing conformation.  The ability of the drug to change the conformation will determine the effect of the drug and tuning this effect will give specific levels of output (i.e., efficacy), initiate only specific types of signaling (i.e., bias) or both.  Tuning of these GPCRs requires studies on how drugs causes changes within the GPCR and how those changes affect the interaction between the GPCR and proteins in the cell.  The duration that a GPCR stays in a particular conformation is known as a dwell time and these dwell times are estimated to occur within 1-10 milliseconds.  To measure these dwell times and characterize their relationship to bias and efficacy, we have developed several types of ultra-fast microscopes (i.e., capable of measuring at speeds of < 1 ms) to monitor the conformational changes of GPCR with drugs of various efficacies and bias.  For the first time, this access will inform on how the dynamics within a GPCR tunes signaling.  In addition, the GPCR is diffusing at the cell membrane where it translates the information from a drug on the outside of the cell to a protein on the inside of the cell.  The interaction between the GPCR and intracellular protein is also defined by the membrane topology and the pools of proteins available for interaction.  To measure these, we have developed methods to simultaneously track or reconstitute multiple proteins to understand how their interactions leads to efficacy and bias.  Overall, we will be able to assess how microscopic events within a GPCR occurring within a defined region of a cell membrane leads to a particular cell response and use this as a determinant to tune drug responses.

Participants

Lead Researcher(s): 

Sponsors: Brian Kobilka (Molecular and Cellular Physiology) and Steven Chu (Physics and Molecular and Cellular Physiology)

Funding Type: 
Postdoctoral Fellowship
Round: 
4
Award Year: 
2018