G protein-linked receptors

Question 1

What is the structure of heterotrimeric G proteins?

Answer 1

Contain three subunits (alpha, beta and gamma). G-alpha and G-gamma are tightly bound to each other while G-alpha dissociates and associates with G-beta-gamma subunit.

Question 1

What is the structure of G protein-coupled receptors?

Answer 1

• Seven transmembrane alpha-helical regions

Question 2

What happens when a hormone binds a GPCR?

Answer 2

• Binding of hormone induces a conformational change in receptor
• Activated receptor binds to Galpha subunit
• Activated receptor causes conformational change in Galpha, triggering dissociation of GDP
• Binding of GTP to Galpha triggers dissociation of Galpha both from the receptor and from Gbeta-gamma
• Hormone dissociates from receptor; Galpha binds to effector, activating it
• Hydrolysis of GTP to GDP causes Galpha to dissociate from effector and reassociate with Gbeta-gamma

Question 3

Explain the fight-or-flight response.

Answer 3

Mediated by epinephrine binding to GPCRs that modulate adenylyl cyclase (effector protein). Activated adenylyl cyclase synthesizes cAMP, which activates protein kinase A.
PKA stimulates glycogen breakdown and inhibits glycogen synthesis:
- Promoter or enhancer segments regulated by PKA contains cAMP-response element (CRE); PKA subunits phosphorylate CREB protein which binds to CRE-containing targets, stimulating gene transcription

Question 4

What are two ways the fight or flight response can be terminated.

Answer 4

1. Termination by Galpha-GTP being hydrozyled or cAMP phosphodiesterase hydrolyzing cAMP to AMP
2. Desentization/adaptation where constant ligand stimulation results in Galpha-GDP binding less well to GPCR. At some point, beta-arrestin can heavily phosphorylate GPCR, blocking Galpha binding, causing endocytosis of GPCR

Question 5

Explain the vision pathway mechanism.

Answer 5

• Photon absorption generates activated rhodopsin, a single photon produces a measurable response, thanks to signal amplification
• Active rhodopsin binds inactive GDP-bound G-protein, which exchanging the GDP for GTP
• GTP-bound Galpha activates PDE, causing the hydrolyzation of cGMP to GMP
• The resulting decrease in cytosolic cGMP leads cGMP to dissociate from the cation channels in the plasma membrane and causes these channels to close
• This causes a hypolarization of the plasma membrane and neurotransmitter is reduced
• Termination of the signal from light-activated rhodopsin by rhodopsin phosphorylation and binding of arrestin
• Rhodopsin kinase phosphorylates rhodopsin, making it less able to bind and activate G-protein
• Arrestin binding to rhodopsin speeds up the inactivation process by preventing rhodopsin to interact with the G-protein

Question 6

What is rhodopsin?

Answer 6

GPCR that is activated at photon absorption.

Question 7

In a dark-adapted state, cGMP-gated ion channels are closed and there is a high cytosolic cGMP. True or false?

Answer 7

False, channels are open. There is high cytosolic cGMP because PDE is inactive.

Question 8

How does the IP3/DAG pathway work?

Answer 8

• Opening of endoplasmic reticulum Ca2+ channels is triggered by ligand binding to GPCRs, leading to activation of phospholipase C
• Cleavage of PI(4,5)P2 by phospholipase C yields IP3 and DAG. IP3 opens IP3-gated Ca2+ channel in the membrane of the endoplasmic reticulum
• Ca2+ in the ER lumen are released into the cytosol, leading to an increase of cytosolic Ca2+

Question 9

What happens when calmudolin is bound to four calcium ions?

Answer 9

• Calmodulin acts as a calcium sensor. When it is bound to four calcium ions, its conformation changes.
• Activated CaM can bind to CaM-Kinase II, partially activating it.
• CaM-Kinase II auto-phosphorylation leads to its full activation; this auto-phosphorylation can occur in the absence of calcium.
• Once auto-phosphorylated, CaM-Kinase II can phosphorylate and activate other CaM-Kinase II complexes

Question 10

What is IP3?

Answer 10

IP3 is a second messenger molecule involved in the phosphoinositide signaling pathway.

Question 11

In light-adapted rod cells, where would expect to see the most arrestin?

Answer 11

In the outer segments where rhodopsin is contained. Arrestin plays a role in desentizing the pathway by binding the phosphorylated rhodopsin and preventing further activation.