Physiology 3140A Lecture Notes - Lecture 18: Heterotrimeric G Protein, Protein Kinase, G Protein-Coupled Receptor Kinase
2 May 2018
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Cell Physiology Lecture 18
GPCR 2
October 30 2017
- What happens when the ligand binds to the receptor?
o Not just the matter of the ligand binding the receptor, signalling taking place, then resetting
- Cells are good at adapting to the amount of receptor they need at the cell surface
- ½ the drugs prescribed are ligands that bind to GPCRs
o Driver for pharmaceutical field
- Important in how cells receive information and transfuse information to the cell to be able to medite
information to the cell, changes to cell beahaviour, etc.
- Regulation of GPCR involves 1) desensitization, 2) down regulation
- Desensitization: Decline in response to continuous or repeated application of agonist (ligand)
o Mechanism for protecting cells against receptor overstimulation
o Purpose of desensitization: receptor can no longer signal to the cell
o Ligand binds to receptor signal gets sent into the cell by second messenger molecule; this
process can be turned off b/c the ligands dissociates or other processes that occur to prevent
the receptor from signalling again
▪ Ligand bound to receptors initiate a process and send a signal cell modifies its
behaviour (cytoskeleton change shape, metabolism, secreted other molecules)
once this happen the cell may not want to do it again (refractory period)
▪ This is mediated by desensitization
o If there is a lot of ligand floating outside the cell, but the cell has already done its function;
does not want to do it again
▪ A way for the receptors to no longer respond to the ligand
o Mechanism for preventing the receptor from being over-stimulated
o RECEPTOR NO LONGER SENSITIVE TO LIGAND OR AGANOST AROUND THE CELL
- Down-regulation: Persistent loss of receptors after long-term exposure to agonist
o Occurs during long-term administration of receptor agonist (tolerance)
▪ Receptor lose their ability to respond; ligand binds and no response
▪ Becomes tolerant to the ligand being present
o Receptors degraded (lysosomes)
▪ Receptor taken into the cell and degraded
o Decreased receptor synthesis: gene transcription/post-transcriptional events
▪ Decreased # of receptors through degradation and not getting replenishment of the
receptors; decrease in synthesis
o PROCESS OF HOW DESENTIZATION OCCURS
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GPCR desensitization
- Receptor Inactivation: exposure of GPCRs to agonists often results in a rapid attenuation of receptor
responsiveness
- Ligand binds to the receptor leading to the activation of the receptor
o Get a conformation change in the shape of the receptor
- Receptor can now couple to the G –protein
o Conformational rearrangement in the receptor; receptors affinity to binding the G protein
increases
- G protein binds; get GEF changing GDP to GTP
- Alpha subunit + beta-gamma subunit dissociates
o Have different effectors that they bind to
o Often is it the alpha subunit, but the beta-gamma subunit can also mediate physiological
impacts in the cell
o E.g. Alpha S binds to adenyl cyclase + cause activation
▪ 2nd messenger cAMP is now made
- Imagine that there is a lot of the ligand, now go into desentization mode
- Desensitization:
o Uncouple of the G protein + no longer able to re-couple to the receptor
o Some mechanisms that can mediate this over a time zone:
▪ Phosphorylation – negative impact to desensitization + G protein uncoupling
• RAPID
▪ Sequestration – receptor gets taken into the cell, therefore no longer at the surface
of the cell (can no longer see the ligand)
• LONGER TERM
▪ Down regulation – degradation of the receptor, etc.
• LONGEST TERM
- Receptor Inactivation
o Exposure of GPCRs to agonists results in rapid attenuation (loss) of receptor responsiveness
o Process, termed desensitization, can be achieved by four mechanisms
- Inactivation takes place when there is a lot of ligand around
- Cells have adapted to the different processes; determined by the amount of ligand around the cell
(large or smaller concentration of ligand)
o If small amount of ligand and the cell has already done it’s job is a different mechanism
compared to when there is large amount of ligand (has to do something more drastic)
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find more resources at oneclass.com
1. Phosphorylation:
- GPCRs are covalently modified by phosphorylation of intracellular serine residues (serine,
threonine, tyrosine) by protein kinases resulting in inability of receptor to couple to heterotrimeric G
protein and stimulate the exchange of GDP for GTP on the -subunit
o Receptor can no longer couple to the G protein (alpha subunit)
o Receptor is phosphorylated – in a conformation that has a low binding affinity to the alpha
subunit
o After the ligand is bound to the receptor, phosphorylation occurs – the affinity for the alpha
subunit is low
▪ THEY UNCOUPLE
- Note: a lot of the signaling processes are highly dependent on the conformation of the individual
proteins involved – conformation of the protein will dictate their binding affinity to other proteins
o Normally, the ligand binds to the GPCR – causes a conformation change in the receptor
▪ Affinity the receptor has for the alpha subunit of the G protein is high
o If the ligand is NOT bound and the conformation is not change, it has a low binding infinity
o Change in conformation in receptor + change in affinity receptor has for the ligand = WHAT
DICTATES IT
- Phosphorylation is most rapid means by which GPCR desensitization can be achieved
- GPCRs phosphorylated within seconds to minutes following ligand binding
- Mediated by:
o 1) Second messenger-dependent protein kinase [cAMP-dependent protein kinase
(PKA) and protein kinase C (PKC)]
▪ When there is a lower concentration of ligand!
▪ Normal activation of the receptor, normal activation of signaling pathways BUT
those signaling pathways feed back to the receptor, you did your job – do not do it
anymore
o 2) Receptor-specific G protein-coupled receptor kinase (GRKs)
▪ When there is a higher concentration of ligand!
▪ Recruits another form of kinases
▪ GRK only come into action when needed as a back up system
2) Internalization:
- GPCRs can be temporarily removed into interior of cell so they are no longer available at cell surface
for ligand binding and unable to interact with heterotrimeric G protein
- Phosphorylation may not be enough – the cell may want to decrease the amount of receptors on the
cell surface
- Note: receptors that go into the cell can cycle back to the surface of the cell
- There can be signals that cause recycling of some of the receptors back to the cell surface
o Ligand binds + send signals into the cell cell behaviour change now the cell doesn’t
want it to happen for a while pulls in the receptor there is a situation where you want
the cell change again receptor goes back to the cell surface
find more resources at oneclass.com
find more resources at oneclass.com
Document Summary
What happens when the ligand binds to the receptor: not just the matter of the ligand binding the receptor, signalling taking place, then resetting. Cells are good at adapting to the amount of receptor they need at the cell surface. The drugs prescribed are ligands that bind to gpcrs: driver for pharmaceutical field. Important in how cells receive information and transfuse information to the cell to be able to medite information to the cell, changes to cell beahaviour, etc. Regulation of gpcr involves 1) desensitization, 2) down regulation. Receptor inactivation: exposure of gpcrs to agonists often results in a rapid attenuation of receptor responsiveness. Ligand binds to the receptor leading to the activation of the receptor: get a conformation change in the shape of the receptor. Receptor can now couple to the g protein increases: conformational rearrangement in the receptor; receptors affinity to binding the g protein. G protein binds; get gef changing gdp to gtp.
