Signal Transduction II

Question 1

What does an agonist do?

Answer 1

It mimics the physiological ligand- it turns on a receptor

Question 2

Synonyms for GCPR

Answer 2

heptahelical, serpentine (due to topography in the membrane)

Question 3

How many transmembrane segments do GCPR have?

Answer 3

7 (sit as a helical bundle)

Question 4

What sits down in the middle of a GCPR bundle?

Answer 4

catacholamine

Question 5

G proteins that interact with GCPR are of what conformation?

Answer 5

triheteromeric

Question 6

What does triheteromeric mean?

Answer 6

Three subunits that are not identical (a, B, y)

Question 7

In the basal state, what is bound to the a subunit of G proteins?

Answer 7

GDP

Question 8

In the basal state, are the a,b, and y subunits complexed together?

Answer 8

Yes

Question 9

Which subunit has intrinsic GTPase activity?

Answer 9

a

Question 10

Which subunit has lipid modification?

Answer 10

a and y

Question 11

Is the y subunit hydrophobic or hydrophilic?

Answer 11

hydrophobic

Question 12

What do lipid modifications do on G protein subunits?

Answer 12

tether them to the membrane (thus, only a and y are tethered to the membrane because b does not have lipid modification)

Question 13

How does GCPR signal transduction begin?

Answer 13

An agonist comes in and binds to a receptor, promotes a conformational change in the receptor, and begins to interact with the G protein

Question 14

What is the rate limiting step in GCPR signal transduction?

Answer 14

GDP release from the a subunit

Question 15

When is GDP released from the a subunit?

Answer 15

When the GCPR binds to the G protein

Question 16

What happens after GDP is released from the a subunit?

Answer 16

GTP (at high conc in the cell) and immediately binds to the empty GDP binding site on the a subunit- this process is unidirectional

Question 17

Binding of GTP to the a subunit causes what?

Answer 17

separation of the a subunit from the B/y subunit - this activates both subunits (a and By)

Question 18

What does the a subunit do once its activated?

Answer 18

it can now act upon a target protein or effector and activate it

Question 19

What causes inactivation of the a subunit?

Answer 19

GTPase activity on the a subunit auto-hydrolyzes the GTP to GDP, causing dissociation of the a subunit.

NOTE: The a subunit then re-combines with the B/y subunit to ‘reset’ the process

Question 20

Do all GCPR work the same?

Answer 20

Yes. Same steps

Question 21

Do adrenaline, glucagon, and adrenocorticotropic hormone work by cAMP transduction?

Answer 21

Yes.

Question 22

Does cAMP diffusion act quickly?

Answer 22

Yes. 100x increase in cAMP in some tissue in as little as 20 sec.

Question 23

When are secondary messengers like cAMP release/synthesized?

Answer 23

When the a subunit becomes activated and binds with an effector protein. The effector protein is responsible for synthesizing/releasing cAMP, not the activated a subunit

Question 24

What synthesizes cAMP? From what substrate?

Answer 24

adenylyl cyclase. from ATP

tightly regulated

Question 25

What breaks down cAMP? Into what?

Answer 25

cyclic AMP phosphodiesterase; into AMP tightly regulated

Question 26

Does the cell want high levels of cAMP?

Answer 26

No, if you force the cell to maintain high levels of cAMP (i.e. block cyclic AMP phosphodiesterase), it will die

Question 27

What are the main steps of adenylyl cyclase activation?

Answer 27

1) normal steps of activation of G protein 2) binding of activated G protein with adenylyl cyclase (thus, activating it) 3) adenylyl cyclase makes cAMP from ATP 4) cAMP activates PKA (protein kinase A) -PKA has many targets in the cell cytoplasm and nucleus

Question 28

What kind of kinase is PKA?

Answer 28

serine/threonine (only phosphorylates on serine or threonine in substrates)

Question 29

How can cAMP affect glycogen breakdown in the cell?

Answer 29

activated PKA (from cAMP) uses ATP hydrolysis to activate phosphorylase kinase. Activated phosphorylase kinase uses ATP to activate glycogen phosphorylase, which causes glycogen breakdown 'kinase cascade'

Question 30

How can cAMP affects gene expression?

Answer 30

activated PKA goes through the nucleopore and activates/phosphorylates a transcription factor, thus up regulating target gene expression

Question 31

How does GPCR regulation of ion channels using B subunits work?

Answer 31

acetylcholine binds to GCPR (skip ahead a couple of steps) the activated B/y subunit complex (which at this point is dissociated from the a subunit due to GTP binding to the a subunit) binds to the K+ channel and opens it inactivation comes when GTP on a subunit is hydrolyzed to GDP, causing re-assembly of subunits to inactive form

Question 32

Phospholipase C breaks which 'head' region off phospholipids?

Answer 32

PIP2- Phosphatidylinositol 4,5, biphosphate

Question 33

fatty acid chain breakdown by Phospholipase C makes what two products?

Answer 33

diacylglyercol and IP3

Question 34

What is diacylglycerol?

Answer 34

The remnant body of the fatty acid chain that remains embedded in the cell membrane following cleavage of the head by phospholipase C (activated PKC)

Question 35

What is IP3?

Answer 35

The 'head' region of the fatty acid cleaved by phospholipase C

Question 36

What does IP3 do?

Answer 36

It binds to receptors on the membrane of the ER and causes release of Ca2+ stores from inside the ER to the cell cytoplasm

Question 37

What does the Ca2+ do once released from the ER by IP3?

Answer 37

Three things: 1) directly binds to some enzymes and activates them 2) activates calmodulin-a Calcium sensor 3 helps in activation of PKCs

Question 38

How does Ca2+ affect calmodulin and what is the action of calmodulin?

Answer 38

Calmodulin is originally sitting in the cell doing nothing until Ca2+ levels rise. In the absense of calcium, calmodulin sits in an almost linear shape. When Ca2+ binds to it, it reconforms its structure to wrap around any calmodulin target (must bind Ca2+ in all four of calcium binding sites)- once it has those holes filled, it wraps around target Once calmodulin binds to target enzyme, it releases the inhibitory domain of enzyme from catalytic domain. The enzyme can then auto-phosphorylate to fully activate itself. Calmodulin then falls off if Ca2+ levels drop, leaving an activated, Ca2+ independent enzyme

Question 39

Does calmodulin have enzymatic activity?

Answer 39

No. it is just a Ca2+ sensor that, when activated by Ca2+, can affect the activity of target enzymes in the cell

Question 40

PKC require binding of what for activation?

Answer 40

Ca2+ and diacylglycerol. Activation of PKC is another consequence of IP3 releasing large amounts of Ca2+ from ER