S2: Protein Breakdown and Urea Formation

Question 1

What is gluconeogenesis?

Answer 1

Formation of glucose from non carbon sources.

Question 2

What is positive and negative nitrogen balance?

Answer 2

We intake proteins through our diet, there are no specific protein stores in our body.

Proteins are either structural or functional and excess protein is broken down and excreted so there should be a balance between input and output. The amino acids can be used to make new protein e.g. muscle fibres, enzymes (structural or functional). The nitrogen is removed in the liver through formation of urea.
Nitrogen free intermediates can then be metabolised into Glucose, Ketone bodies and CO2 and H2O.

Positive nitrogen balance:

• Amount of protein/AA we retain exceeds the amount that is broken down and excreted.
• This is a normal process and occurs in things like growth

Negative nitrogen balance:
- Input is superseded by breakdown

Question 3

What are some reasons for positive nitrogen balance?

Answer 3

• Growth in small children
• When someone is pregnant - they will be taking in and laying down more protein
• Takes place in response to excersize where there is tissue hypertrophy as well as a response to anabolic hormones

In positive nitrogen balance, more of the AAs in the AA pool are being converted into body protein and less body protein is being broken down or excreted.

Question 4

What are some reasons for negative nitrogen balance?

Answer 4

• May be caused due to protein deficiency
• e.g. wasting disease, burns and trauma
• It could also be in response to catabolic hormones, or lack of anabolic ones e.g. diabetes

Negative nitrogen balance can cause someone to lose body protein mass.

Question 5

What are the two steps in the metabolism of amino acids?

Answer 5

The first step is the breaking down of protein/polypeptide via peptidases into its constituent amino acids.

Normal body protein metabolism means dealing with amino acids in two parts:

1. Dealing with carbon skeleton
2. The Nitrogen

Question 6

Describe the metabolism of Amino acids (carbon skeleton and removal of nitrogen)

Answer 6

Carbon Skeleton: The carbon skeleton can be used for energy metabolism or biosynthesis.

Removal of Nitrogen:

Nitrogen is toxic (adverse effect on neuronal cells) so has to be removed safely. Individuals who cannot produce urea often die in infancy.
In mammals, the nitrogen is converted to the non-toxic neutral compound urea and excreted in the urine.

The process by which the amino acid nitrogen is transferred to urea is a three step process:

1. Transamination
2. Formation of ammonia 
3. Formation of urea.

Question 7

What is the equation for the formation of ammonia from amino acid?

Answer 7

Amino Acid –> a-keto acid + NH3

Nitrogen removed from AA forming a-keto acid and ammonia (toxic). This ammonia usually exists as ammonium ions.

Question 8

Can urea be formed in muscle?

Answer 8

No as the enzyme to do this is not present.

However, the carbon skeleton can be obtained and used for energy.

Question 9

What is transanimation?

Answer 9

The nitrogen as part of the a-amino group is transferred to an a-keto acid to become a new amino acid

Question 10

Name 3 a-keto acids and their uses

Answer 10

TCA cycle intermediates:

• a-ketoglutarate
• Oxaloacetate
• Pyruvate

α-ketoglutarate, pyruvate and oxaloacetate can be oxidised or converted to make glucose (supplementing gluconeogenesis).

Question 11

What enzyme carries out transanimation?

What are the two most important ones?

Answer 11

The enzymes that do this are transaminases/aminotransferase, there are quite a lot of different types of transaminases.

The most important are the alanine (ALT) and aspartate (AST) transaminases

Question 12

What are the reactions that alanine transaminase (ALT) and aspartate transaminase (AST) catalyse?

What amino acid from both reaction is produced?

Answer 12

ALT -
Alanine + a-ketoglutarate –> pyruvate + glutamate

AST -
Aspartate + a-ketoglutarate –> oxaloacetate + glutamate

The amino acid glutamate is produced in both reactions. These reacts are in equilibrium.

Question 13

Why is glutamate important?

Answer 13

Glutamate is a way the body can transport potentially toxic N

Question 14

How can transaminases be used diagnostically?

Answer 14

The transaminases are primarily liver enzymes so can be used diagnostically,
and high levels of AST and ALT in the blood are indicative of liver damage (as they normally shouldn’t be found in plasma)

Question 15

What does alanine need to work?

Answer 15

Vitamin B6

Question 16

What happens to the glutamate made during transamination?

Answer 16

• Glutamate can release the ammonia by action of a second enzyme, glutamate dehydrogenase that is present in the mitochondrial matrix (transamination occurs in cytosol).
• It will yield back α-ketoglutarate.
• NAD or NADP can be used, however it is usual for NAD to be used for degradation and NADPH for synthesis.
  This process is called oxidative deamination.

Glutamate is a very useful molecule because it is freely interchangeable with the α-keto acids as well as the ability to donate and accept ammonium ions.

Question 17

Why is transamination to glutamate and oxidative deamination back to a-ketoglutarate important?

Answer 17

The reason it is very important is because it allows conversion of many amino acids from their original state into glutamate, which can be transported (note it is not often transported as glutamate) and then re-converted back into something the body can use for energy (or transamination again!) while re-synthesising the ammonia which can be fed into the urea cycle.

Question 18

Explain how free ammonia can be eliminated

Answer 18

Glutamate formed as a result of transamination can be used to transport another nitrogen which is part of an ammonium ion.

Glutamate + NH4+ + ATP Glutamine + ADP

Glutamine synthase is the enzyme

• Glutamine is like another transport molecule, a way in which the body can transport the potentially toxic N to the liver.
• This is a reaction that often takes place in the periphery, glutamine synthase is widely distributed, esp. in blood vessels with a lot of protein breakdown including those blood vessels of the liver itself.

The reaction goes both ways so we can resynthesise the glutamate from the glutamine.

Question 19

Describe the urea cycle

Answer 19

• The urea cycle is a metabolic pathway that is the means for excreting nitrogen
• It is restricted in its distribution, being mostly in the liver.
• It does not exist in the muscle
• It takes place in the mitochondria and cytoplasm of heptocytes
• The urea cycle uses as its substrates bicarbonate, aspartate and ammonium ions (released from glutamine or glutamate)

Question 20

Where does bicarbonate as a substrate for urea cycle come from?

Answer 20

The bicarbonate comes from the breakdown of the carbon skeleton (i.e. CO2 as a by product of metabolising the carbon skeleton is formed into bicarbonate which is then used to obtain the CO2 needed)

Question 21

Describe structure of urea

Answer 21

Here is urea, it has two nitrogen atoms. One of them is donated from aspartate, while the other comes from glutamine/glutamate.
The carbon C=O comes from the carbon skeleton, through using CO2 that has been produced from its breakdown.
Hence, the detrimental products of amino acid degradation can be used to combine to form urea, a non-toxic, soluble compound that can be readily excreted.

Question 22

How is urea cycle and TCA cycle linked?

Answer 22

The CO2 comes from the bicarbonate and reacts with the ammonium ion that has come from glutamine/glutamate (formed by transamination of α-ketoglutarate and α-amino acid). They form Carbamoyl phosphate.

The starting point could be thought to be Aspartate, formed by the transamination of α-amino acids, when reacting oxaloacetate (another type of keto-acid) with an α-amino acid. And the formation of the Carbamoyl phosphate

Question 23

Can muscle form urea?

Answer 23

No its doesn’t have the enzymes

Question 24

What type of amino acid does muscle break down during prolonged excersize or starvation?

Answer 24

Branched amino acids are used for this energy (branches e.g. leucine)

Question 25

What two routes are the remaining amino acids (ones not broken down in muscle) dealt with in?

Answer 25

• Nitrogen is transferred to alanine via glutamate and pyruvate
• Circulating/Intracellular Glutamate can be made into glutamine (return to liver)

Question 26

Glucose alanine cycle

Answer 26

The muscle can export alanine, it is one of the major exports of muscle that is actively being broken down (due to exercise or starvation).

• Branched AA are taken, the carbon skeleton is used for energy production. Then the NH4 can be used to convert to pyruvate to Alanine.
• Alanine is then exported into the blood and travels to the liver.
• The alanine is then converted to glutamate via transamination (reacting with α-ketogluterate) also producing a pyruvate.
  The pyruvate can enter the gluconeogenic pathway to form glucose, the glucose can be transported in the blood back to the muscle where it can be used for energy.
  The glutamate will then be used along with the CO2 generated to produce urea.

So there is this cycle between the two important tissues.
s

Question 27

What are the two types of amino acids and what do they do?

Answer 27

1. Ketogenic amino acids
   - They will form ketone bodies
2. Glucogenic amino acids
   - Used by the liver to produce glucose

Some amino acids fall into both categories.
Amino acids can feed into different parts of this main carbohydrate pathway.
This occurs in the liver, where most of the gluconeogenic pathway takes place. Amino acids broken down can be converted into intermediates of the TCA cycle. This is good because we can convert this back to glucose.