Liver physiology: Hepatic protein metabolism and amino acids in nitrogen balance

Question 1

What is the main source of nitrogen from our diet?

Answer 1

The main source of nitrogen is from dietary protein, the main loss from the gut and kidneys (as urea).

Question 2

How does the body maintain a pool of free amino acids?

Answer 2

The body maintains a pool of free amino acids in the blood. In a fed state, the net contributor is the diet; in a fasting state it is bodily protein (of which 80% is in skeletal muscle).

Question 3

How can amino acids be used?

Answer 3

Amino acids can be used in protein (and other nitrogenous product) synthesis, or lose the amino group to leave a carbon backbone for glucose synthesis and energy metabolism.

Question 4

What does an amino acid look like (in terms of chemical formula)?

Answer 4

• Amino group (NH2)
• Side chain (R)
• Carboxylic acid group (COOH)

Question 5

What is the idea of nitrogen balance?

Answer 5

The amount that is excreted (in form of urea) and the amount of nitrogen intake
- we want to be in equilibrium

Question 6

Why would someone be in a positive or negative nitrogen balance?

Answer 6

If they are in a positive balance they are gaining proteins and if they are in a negative balance they are losing proteins

Question 7

How are free amino acids split in the body?

Answer 7

• Free pool in blood
• Proteins in the body (~10 kg) and other nitrogenous products
• Metabolic precursors – loss of the amino group leaving the carbon backbone used in glycolysis and TCA cycle intermediates
• No amino acid is stored as they always have a function

Question 8

Who may have a positive nitrogen balance?

Answer 8

• Pregnant women
• Lactation
• Bodybuilder

Question 9

Who may have a negative nitrogen balance?

Answer 9

• Protein malnutrition
• Severe illness/ sepsis
• Essential amino acid deficiency

Question 10

What is kwashiorkor?

Answer 10

They have the adequate amount of calories but an inadequate amount of protein
- Oedema of stomach, fatty liver

Question 11

How are amino acids metabolised in the fed state (after you have eaten first few hours)?

Answer 11

• Dietary protein digested into amino acids in the gut
• Then transported to the liver and can be used to be made into proteins or other essential nitrogen-containing compounds
• The excess amino acids have their amino group removed and carbolic backbones used as metabolic substrates to form glucose and glycogen etc…

Question 12

How are dietary proteins broken down into denatured proteins?

Answer 12

• By HCl and pepsin
• In the stomach

Question 13

How are denatured proteins broken down into Oligopeptides and Amino Acids?

Answer 13

• By Chymotrypsin(ogen), Trypsin(ogen) and Aminopeptidase
• In the small intestine

Question 14

How are Oligopeptides and Amino Acids absorbed into the bloodstream?

Answer 14

• by enterocyte peptidases in the enterocytes (intestinal absorptive cells)
• Amino acids are absorbed into the lumen with Na+ and then the amino acid is actively transported with ATP into the portal vein

Question 15

What are glucogenic AA?

Answer 15

Carbon backbone used in gluconeogenesis or TCA cycle intermediates

Question 16

What are ketogenic AA?

Answer 16

Carbon backbone produces Acetyl CoA/ Acetoacetyl CoA
(Only LEUCINE and LYSINE are solely ketogenic)

Question 17

What are some essential AA?

Answer 17

Phenylalanine
Valine
Leucine

Question 18

What are some conditionally essential AA?

Answer 18

Arginine
Cysteine
Glycine

Question 19

What are some non-essential AA?

Answer 19

Alanine
Glutamate
Aspartate

Question 20

When are amino acids essential?

Answer 20

If they can not be synthesised de novo in the body

Question 21

Why is Albumin so important?

Answer 21

• Made in the liver
• Binding and transport (carrier protein)
• Maintenance of colloid osmotic pressure
• Free radicals
• Anticoagulant effects

Question 22

Describe the amino acid degradation 1: Fate of Carbon backbone

Answer 22

• This is when amino acids have been absorbed and enter the free pool of amino acids
• Because they can’t be stored something needs to happen with it
• Amino group (NH2) is removed and forms alpha-keto acids which is the carbon backbone
• A lot of these alpha-keto acids are used in TCA (e.g. remove amino group from alanine gives pyruvate)

Question 23

What is transamination?

Answer 23

• Turning an amino acid into an intermediate in the TCA cycle
• Catalysed by an amino transferase e.g. pyridoxal phosphate (PLP) derived from vit B6
• Taking an amine group from an amino acid to an alpha-ketoacid
• Turns it into an amino acid and becomes an alpha-ketoacid itself

Question 24

How does transamination of alanine work?

Answer 24

• Alpha-ketoglutarate is a universal acceptor of amine groups
• Alanine uses the enzyme alanine aminotransferase (ALT) – forms pyruvate and glutamate (the normal knew AA)

Question 25

Describe the Degradation of Amino Acids 2: Fate of Nitrogen

Answer 25

• Glutamate reacts with NADP + H20 (under the influence of Glutamate Dehydrogenase)
• forms ammonium (NH4+)
• The carbon backbone is kept and forms an alpha-ketoacid which can be recycled to form more glutamate (under transamination)
• Ammonia then enters the urea cycle and excreted as urea

Question 26

How are amino acids formed in the fasting state?

Answer 26

• Protein in skeletal muscle broken down into AA’s which can enter TCA cycle and form ATP, alanine and other AA
• Can also be used for gluconeogenesis to form glucose for energy
• Alanine returns to liver where turned back into pyruvate to be turned into glucose

Question 27

Why do proteins have different lifespans?

Answer 27

• Become obsolete
• Faulty and need to be removed
• Flexible system required to meet
  energy requirements

Question 28

What is the mechanism of degradation with proteasomes?

Answer 28

• Small protein (ubiquitin)
• Through the action of 3 enzymes it forms an isopeptide bond with multiple lysine residues
• This signals to proteasomes that the protein needs to be degraded
• Produces a signal. This is stronger when >4 ubiquitin proteins together

Question 29

What are the enzymes involved with proteasome degradation?

Answer 29

E1 – ubiquitin-activating enzyme
E2 – ubiquitin-conjugating enzyme
E3 – ubiquitin-protein ligase

Question 30

What is a proteasome?

Answer 30

Once protein marked with ubiquitin the actual job with degrading the protein is the proteasome
- It consists of the proteasome (contains ATP dependent hydrolyses which break down the protein) and caps either end which regulates which proteins may enter the proteasome for destruction

Question 31

What is the N-terminal rule?

Answer 31

• N-terminal determines the proteins half-life – depends on the amino acid present
• PEST sequences promote ubiquitination (proline, glutamate, serine, threonine)
• Cyclin destruction box promotes

Question 32

What is the lysosomal mechanism of degradation?

Answer 32

Lysosomal – non-selective “ball of death”
- Digests proteins, carbohydrates and fats
- Proteolytic enzymes within lysosome separated from cytosolic components
- Main process is autophagy

Question 33

What are the methods of degradation? (autophagy)

Answer 33

MACROAUTOPHAGY -non-selective:
- ER derived autophagisomes engulf cytosolic proteins/aggregates organelles. Lysosome fuses with this to initiate proteolysis.

MICROAUTOPHAGY-non-selective
- Invaginations of lysosomal membrane engulf proteins/organelles.

CHAPERONE-MEDIATED AUTOPHAGY-selective
- Chaperone protein hsc70, in cytosol and intralysosomal, accompany specific cytosolic proteins in response to stressors (fasting/ oxidative stress etc).

ENDOCYTOSIS/PHAGOCYTOSIS
- Extracellular substances.