Regulation of fatty acid biosynthesis

Question 1

The catalytic activity of what affects the chain length of the fatty acid product

Answer 1

1. Thioesterase

Question 2

What are 4 factors that affect the rate of fatty acid biosynthesis

Answer 2

1. Hydroxytricarboxylic acid levels (e.g. citrate)
2. Long chain acyl-CoA levels
3. Phosphorylation status
4. Enzyme synthesis/degradation

Question 3

Does citrate inhibit or assist fatty acid biosynthesis

Answer 3

1. High levels of citrate increase activity

Question 4

How does citrate increase activity

Answer 4

1. Citrate is a precursor of Acetyl-CoA, the substrate of the enzyme
2. High levels of citrate increase activity
3. Citrate levels are only high when there is excess energy
4. Citrate is a positive feed-forward activator
5. Citrate conc. in cytosol, estimated at 0.3 - 1.9mM
6. Can do enzyme assay- find characteristics of enzyme by varying concentration with ACC
7. Close to in vitro conc determined for half maximal activity of ACC

Question 5

How do you know that changes of citrate concentration is going to have an effect in vitro

Answer 5

1. Enzyme assays in solution
2. Vary substrate concentration and vary citrate concentration
3. In vitro with ACC- half maximal activity of ACC with citrate concentration you would get in cytosol
4. Show changes in cytosolic concentration is going to have an effect

Question 6

How do long chain acyl-CoA levels affect fatty acid biosynthesis

Answer 6

1. Most effective inhibitors are saturated 16-20 C acyl CoA

2. Unsaturated fatty acids are much less inhibitory

Question 7

Describe how long chain acyl-CoA inhibits

Answer 7

1. Inhibition by long chain acyl-CoA is competitive with citrate
2. But non-competitive towards the substrates acetyl-CoA, ATP and bicarbonate ions
3. Inhibition constant is 5.5nM- concentration which gives 50% inhibition
4. Not known if cytoplasmic concentration is close to 5.5- not known if they have effect in body
5. Would have to burst membrane to measure cytoplasmic conc.
6. Used different methods of disrupting cells to see if they agree

Question 8

What effect does phosphorylation have on the rate of fatty acid biosynthesis

Answer 8

1. Reduces activity

Question 9

How does phosphorylation reduce the activity

Answer 9

1. Six possible sites for phosphorylation
2. Hydroxyl groups of serine side chains
3. Phosphorylated form of ACC is INACTIVE or heavily reduced activity
4. Excess energy is used to make fatty acids
5. By adding a phosphate group it changes the shape of the enzyme which changes the activity

Question 10

How are phosphate groups removed

Answer 10

1. Phosphate groups can be removed relatively easily- alkaline phosphatase

Question 11

What is the name of enzymes that add a phosphate group to a protein

Answer 11

1. Kinase

Question 12

How does ACC become active and how does citrate/ phosphate affect this

Answer 12

1. Acc polymerises to be active
2. Dimers (inactive) polymers (active)
3. E.g. binding of citrate promotes formation of polymers and binding of phosphate promotes formation of dimers
4. Dimers are still active just a lot less active
5. Vmax can change by up to 20 fold

Question 13

Describe how the shift between ACC dimer to polymer changes with different factors

Answer 13

1. Citrate shifts equilibrium towards polymerisation i.e. activating
2. Acyl-CoA shifts equilibrium towards dimers i.e. inactivating
3. Phosphorylation shifts equilibrium towards dimers i.e. inactivating
4. The extent of activity is a balance of these.
5. However, the predominant factor under most conditions is phosphorylation- can be altered most rapidly- also responds to signals outside the cells

Question 14

What is the long term mechanism of regulation

Answer 14

1. How much protein is present in cell
2. Alteration of biosynthesis of ACC
3. Starved rats only have about a quarter to a fifth of the ACC of normally fed rats
4. The main method is control of the rate of synthesis

Question 15

What is the main stage to control rate of synthesis

Answer 15

1. The main stage of control of synthesis is at the level of transcription
2. Specific transcription factors have been identified, a key one being SREBP1c (sterol-regulatory-element-binding protein 1c)
3. Major control factor for these transcription factors is the level of unesterified fatty acid

Question 16

What hormones are the main regulators of metabolism

Answer 16

1. Insulin and glucagon are the main regulators of metabolism
2. Control carbohydrate, amino acid and fat metabolism

Question 17

How can you control the rate at which ACC becomes phosphorylated

Answer 17

1. AMP-activated protein kinase controls phosphorylation
2. Only get AMP when low levels of energy
3. So AMP-activated protein kinase is only active when there is little of the material (ATP) that you want to make into fatty acids to be stored

Question 18

What activates AMP-activated protein kinase

Answer 18

1. AMP-activated protein kinase is active when attached to a phosphate

Question 19

What controls the phosphorylation of AMP-activated protein kinase

Answer 19

1. The phosphorylation of AMP-activated protein kinase is activated (phosphorylated) by Kinase kinase
2. ATP–>ADP

Question 20

What effect does insulin have

Answer 20

1. Positive effect on dephosphorylation of AMP-activated protein kinase
2. Inactivating it
3. Leaves ACC in active form
4. ACC can convert food which is what activated insulin

Question 21

What effect does glucagon have

Answer 21

1. Stimulates cAMP
2. cAMP activates kinase kinase
3. Results in formation of more phosphorylated ACC so inactivates it

Question 22

What effect does fatty acyl-CoA have

Answer 22

1. Inhibits acetyl-CoA carboxylase
2. Activates kinase kinase
3. Two different ways of inhibiting ACC