Unit 4 - Part 3

What is the #1 carbon fatty acid found on triacylglycerol and phospholipids?

stearate

How is fatty acid entry into mitochondria regulated so that they cannot be broken down under high ATP conditions?

1. fatty acids allowed into cell under low ATP conditions (high AMP)

2. AMPK inhibits ACC (makes malonyl CoA)

3. lack of malonyl CoA allows carnitine transferase to move fatty acids into mitochondria

secondary alcohol

OH- attached to a carbon that is bonded to 2 other carbons and 1 hydrogen

Where does beta oxidation occur?

in the matrix of the mitochondria

What is the first step of beta oxidation?

beta carbon (C3) of acyl CoA donates a hydride to FAD → produces FADH and 2-enoylCoA

How many ATP is acetyl CoA worth?

10 ATP

What happens in beta oxidation once 2-enoylCoA is produced?

the double bond of 2-enoylCoA is hydrated → produces 3-hydroxyacylCoA

Which structure in the beta oxidation pathway has a secondary alcohol?

3-hydroxyacylCoA

What happens in beta oxidation once 3-hydroxyacylCoA is formed?

it is oxidized by NAD → produces NADH and 3-ketoacylCoA

What happens in beta oxidation once 3-ketoacylCoA is formed?

thiolase enzyme makes the keto group from 3-ketoacylCoA attacks the sulfur from a CoA to make a new CoA → produces acyl CoA and acetyl CoA

When are fatty acids made in abundance?

under low glucose conditions

How does glucose/glycogen compare to fatty acids?

• fatty acids are easier to store than glycogen

• fatty acids yield more ATP than glucose

What happens if you run low on 3-carbon compounds for gluconeogenesis?

muscle is broken down to feed alanine into gluconeogenesis

ketone bodies

emergency solution for the brain made by excess acetylCoA when glucose is too low

ketogenesis

process done when glucose is too low and there are too few 3-carbon compounds to feed gluconeogenesis

What are the options for what can be done with acetylCoA made by beta oxidation?

it can feed into the citric acid cycle or ketogenesis

What is the first step of ketogenesis?

thiolase removes CoA from one acetylCoA to combine 2 acetylCoAs together → produces acetoacetylCoA

What happens once acetoacetylCoA is formed in ketogenesis?

acetoacetylCoA is reacted with another acetylCoA and H2O → produces 3-hydroxy-3-methylglutarylCoA by removing a CoA

What happens in ketogenesis once 3-hydroxy-3-methylglutarylCoA is made?

acetylCoA is removed from it → produces acetoacetate

What happens in ketogenesis once acetoacetate is made?

• 90% of the time: beta carbon of acetoacetate is reduced by NADH → produces 3-hydroxybutyrate

• 10% of the time: acetoacetate loses the carboxylate (in the form of CO2), which is chemically favorable without an enzyme → produces acetone

What is the major ketone body?

3-hydroxybutyrate

What is the minor ketone body?

acetone

What does the NAD+ that is formed in the forming of 3-hydroxybutyrate get used for?

recycled back to restore NAD+ used to make 3-keto-acylCoA

Which molecule in the ketogenesis pathway FIRST has a carboxyl group?

3-hydroxy-3-methylglutaryl CoA

What happens to the main ketone body in the CNS?

1. 3-hydroxybutyrate is oxidized via dehydrogenase → produces NADH and acetoacetate

2. CoA transferase combines acetoacetate with succinylCoA → produces succinate and acetoacetylCoA

3. thiolase uses a CoA to cut acetoacetylCoA in half → produces 2 acetylCoAs

How much ATP do you get from 1 main ketone body compared to glucose?

22.5 ATP for ketone body and 32 ATP for glucose

norepinephrine

hormone secreted by adrenal medulla onto the pancreas in response to sympathetic NS low blood glucose

What activates glucagon secretion?

norepinephrine

How does protein kinase A get activated?

1. norepinephrine/epinephrine/glucagon binds to beta adrenergic receptor

2. this activates GEF to eject GDP and pick up GTP

3. When Gas is bound to GTP, it stimulates adenylate cyclase

4. adenylate cyclase picks up ATP → cAMP

5. cAMP binds R subunit and removes it so that the active site of protein kinase A is open

How is cAMP made from ATP?

ribose’s 3’ OH- makes a cyclic bond with 5’ phosphate and P-Pi is biproduct

What enzyme converts cAMP to AMP?

phosphodiesterase

ACTH

hormone produced by anterior pituitary in response to low blood glucose that indicates stress

What activates cortisol release?

ACTH release

cortisol

hormone produced by the adrenal cortex in response to ACTH that activates the production of glucose

What part of the adrenal cortex produces cortisol?

zona fasiculata

pseudosubstrate sequence of R subunit

has a high affinity for the active site of protein kinase A but it lacks the actual amino acid to which you would attach a phosphate

What is protein kinase A also sometimes called?

catalytic subunit

taken up by alpha cells through its liganded GABA-R

chloride

forms a barrier preventing breakdown of triacylglycerol (TAG) within adipose cells

perilipin

acts as a GEF on GalphaS/GDP in response to norepinephrine or epinephrine

beta-2-adrenergic receptor

activates the removal of the separation between triacylglycerols and lipases

PKA

places a pseudosubstrate sequence into the active site of PKA to keep it inhibited

R subunit

hydrolyzes the 3'-phosphoester bond in cAMP in a PIP3-dependent state

PDE3b

released by activated beta cells to hyperpolarize Islet alpha cells

GABA

activated by GalphaS/GTP to produce cAMP

adenylate cyclase

converts TAG to DAG and one free fatty acid within adipose

ATGL

target of PKA that hydrolyzes the C2 acyl chain from diacylglycerol (DAG) in adipose

hormone-sensitive lipase

product of wortmannin-sensitive p110 in response to insulin

PIP3

emulsifies TAGs at the cytoplasm/lipid boundary to feed them to ATGL

CA

target of PKA that activates glucagon secretion in response to norepinephrine

L type calcium channels

activates the removal of the pseudosubstrate sequence from PKA

cAMP

reverses PKA product within adipose cells in response to insulin, inhibiting lipolysis

PP-1

How does cAMP remove the pseudosubstrate from the active site of protein kinase A?

it extends an alpha helix

What is the pseudosubstrate sequence and what purpose does each amino acid serve?

RR (positively charged that interacts with glutamate), G (H+), A (methyl group), I (large, nonpolar, hydrophobic)

What can the first R in the pseudosubstrate sequence be replaced with sometimes?

K but it will be weaker

L type Ca++ channels

allows calcium to flow into the cell following depolarization and binding of protein kinase A to its cytoplasmic binding site and activates ryanodine receptors to open

Protein Kinase A Substrate sequence and their purpose

RR (positively charged to interact with glutamate), X (something small), S/T (has an OH- group), L (non-polar and hydrophobic)

How does L-type calcium channels change the action of myocytes?

epinephrine binds to beta-adrenergic receptor → activates PKA → Ca2+ influx → secretion and increased HR and contraction in myocytes

PDE3b (phosphodiesterase 3b)

converts cAMP back into AMP in response to insulin

What are the two ways that insulin can suppress glucagon secretion?

1. inhibiting L-type Ca2+ channels via PKA

2. inhibiting L-type Ca2+ channels via hyperpolarization

Describe the steps to how insulin suppresses glucagon secretion by inhibiting calcium channels via PKA

1. insulin binds to IR on alpha cells and activates wortmannin-sensitive pathway

2. p110 converts PIP2 to PIP3

3. PDE3b binds to PIP3 through its PH domain and converts cAMP → AMP

4. R subunit comes back and inhibits protein kinase A with pseudosubstrate sequence

5. PKA cannot phosphorylate L-type Ca2+ channels

6. Ca2+ does not release glucagon-filled vesicles from synaptotagmin to be secreted

Describe the steps to how insulin suppresses glucagon secretion by inhibiting calcium channels via hyperpolarization

1. insulin binds to IR on alpha cells and activates wortmannin-sensitive pathway

2. p110 converts PIP2 to PIP3

3. PIP3 activates Akt-PKB

4. Akt-PKB phosphorylates AS160 (can no longer act as a GAP on Rabs)

5. Rab X progresses to the membrane to get GABA into membrane

6. GABA binds to receptor and activates chloride ions into the cell (hyperpolarization)

What does GABA stand for?

gamma-aminobutyric acid

Rab X

guides receptors for GABA to get into membrane

paracrine

alpha and beta cells are side by side and insulin and GABA go from beta → alpha cells

Where are triacylglycerols stored in adipose cells?

lipid droplet

What does glut 4 do in adipose cells?

takes up glucose to convert DHAP → glycerol 3-phosphate (backbone for triacylglycerol)

PP-1

responds to insulin action when Akt-PKB phosphorylates it by removing phosphates from HS lipase and ATGL

VLDL

large lipoproteins going through the bloodstream made by the liver that deliver fat to tissues

chilomicron

large lipoproteins going through the bloodstream coming from the diet that deliver fat to tissues

When are HS lipase and perilipin in their active forms?

when they are phosphorylated

What are the two ways that insulin inhibits lipolysis

1. PP-1 removing phosphates from the proteins that have been phosphorylated by protein kinase A

2. lowering levels of cAMP in adipose cells

Describe the steps of lipolysis?

1. glucagon binds to glucagon receptor which activates Gas

2. Gas activates adenylate cyclase to make cAMP

3. cAMP bonds to the R subunit to activate protein kinase A

4. protein kinase A phosphorylates perilipin

5. perilipin dissolves the barrier between triacylglycerols and adipose triglyceride lipase

6. one at a time, co-activator brings triacylglceride into the active site of ATGL and its #3 acyl chain is removed to form diacyl glycerol

7. hormone-sensitive lipase converts DAG → monoacyl glycerol

8. monoacyl glycerol lipase cleaves glycerol chain to form 3 fatty acids (total) and a glycerol

What does insulin promote?

triacylglycerol storage

What does insulin inhibit?

lipolysis, epinephrine, glucagon, gluconeogenesis

What does epinephrine/glucagon promote?

lipolysis, glycogen breakdown, gluconeogenesis

What does epinephrine/glucagon inhibit?

triacylglycerol storage

Where is epinephrine and norepinephrine produced and by what cells?

in the adrenal medulla by chromaffin cells

What is the ratio for how much norepinephrine vs epinephrine is produced?

30% norepinephrine and 70% epinephrine

a glucocorticoid that raises blood glucose by increasing transcription of gluconeogenesis enzymes

cortisol

What does cortisol promote?

gluconeogenesis and proteolysis

What gland controls secretion of cortisol?

anterior pituitary

What cells secrete cortisol and where?

corticotrope cells in zona fasciculata in adrenal gland

How is gluconeogenesis regulated?

by the transcriptional activation of PEPCK, fructose 1,6 bisphosphatase, and glucose 6-phosphatase

FoxO1

binds to minor groove of DNA to displace H1 and unwind DNA at IRE region

What type of histone is H1?

inter-nucleosomal histones that tie histones together and space them properly

How does insulin regulate Fox01

Fox01 gets phosphorylated by Akt-PKB in response to insulin which inactivates it

response element

short sequence of DNA in the promoter region that binds TFs

What are the three response elements in the regulation of gluconeogenesis?

CRE, GRE, and IRE

CREB

protein that is phosphorylated by PKA following glucagon/epinephrine stimulation and binds to the CRE region

CBP/p300

co-activator that binds to CREB and has multiple domains

What are the domains of CBP

1. KIX

2. histone acetyl-transferase

3. bromo

KIX domain of CBP

recognizes phosphates found on certain transcription factors (CREB)

histone acetyl transferase domain of CBP

allows CBP to attach acetyl groups to lysines on octamer

bromo domain of CBP

binds to acetyl-lysines (bromo domain interactions spread histone acetylation)

How does the CREB, CBP, and CRE play out in regulation of gluconeogenesis?

1. CREB binds to CRE and gets phosphorylated at a serine by protein kinase A

2. the KIX domain of CBP binds to CREB

3. CBP binding to phosphorylated CREB activates acetyl transferase domain

4. the bromo domain of CBP spreads histone acetylation

How does histone acetylation make transcription happen?

it reduces the charge interaction between a positive lysine and negative DNA to weaken the interaction and allow room for TFs to bind

nucleosome

8 histones with 2 wraps of DNA around it

Which part of histones get modified?

amino terminal tail with lysine and arginine

Which types of histones are most important for modifications and why?

H3 and H4 because they have the longest tails