Biochemistry- TCA cycle

Question 1

Why does NAD+ which was reduced in glycolysis to NADH need to be recycled?

Answer 1

There are limited amounts of NAD+ in the cell. We cannot synthesise it ourselves- it derived from a vitamin niacin.
Therefore NADH must be re-oxidised to allow glycolysis to continue.

Question 2

Describe ways in which NAD+ is recycled (i.e. NADH is re-oxidised) in the absence of oxygen, and name the organisms in which these pathways occur.

Answer 2

Alcoholic fermentation: yeast and some other microorganisms can form ethanol from pyruvate in the absence of oxygen by the action of alcohol dehydrogenase. This is a reductive enzyme so requires the input of electrons. These are donated by NADH, which is regenerated to NAD+. Humans cannot form ethanol from pyruvate.

Lactic acid fermentation:
some microorganisms and humans can form lactate from pyruvate. The reaction is catalysed by lactate dehydrogenase which is a reductive enzyme and requires an input of electrons, which are donated by NADH. This then allows glycolysis to continue to occur when there is a lack of oxygen (e.g. in a hardworking muscle cell). In a human, this will not continue for long as there will be a build up of lactic acid which is harmful to the cell and H+ will inhibit glycolytic enzymes.

Question 3

What are the three names for the citric acid cycle?

Answer 3

Citric acid cycle, TCA cycle, Krebs cycle.

Question 4

How many compartments do mitochondria have?

Answer 4

4- the outer membrane, the inner membrane, the intermembrane space and the matrix.

Question 5

Where in the mitochondria does the vast majority of energy metabolism occur?

Answer 5

The matrix

Question 6

What are the invaginations of the inner mitochondrial membrane called?

Answer 6

Cristae

Question 7

Where does the Krebs cycle occur?

Answer 7

In the matrix of the mitochondria

Question 8

What is the substrate for the krebs cycle?

Answer 8

Acetyl coA

Question 9

What happens to pyruvate after it has been formed in glycolysis?

Answer 9

It was formed by glycolysis in the cytoplasm of the cell. It moves into the matrix of the mitochondria via a specific transporter.
There the pyruvate dehydrogenase complex catalyses the oxidative decarboxylation of pyruvate to acetyl coA. One CO2 molecule is released and the remaining 2 carbon atoms end up as the acetyl group in acetyl coA.
This is an oxidation reaction so NADH + H+ is formed.

Question 10

Where does the pyruvate dehydrogenase complex reside?

Answer 10

In the matrix of the mitochondria.

Question 11

How many enzymes make up the pyruvate dehydrogenase complex?

Answer 11

3

Question 12

How many enzymes are involved in the control of the pyruvate dehydrogenase complex?

Answer 12

2: a kinase and a phosphatase in a single polypeptide.

Question 13

How many coenzymes are involved in the action of the pyruvate dehydrogenase complex?
What are they?

Answer 13

5.

They are thiamine, lipoic acid, coenzyme A, FAD and NAD+.

Question 14

Is the reaction catalysed by the pyruvate dehydrogenase complex reversible?

Answer 14

No, it is irreversible. Acetyl coA cannot be converted back to pyruvate.

Question 15

What is the major determinant of glucose oxidation in well oxygenated tissues in vivo?

Answer 15

The activity of PDC

Question 16

How many reactions make up the TCA cycle?

Answer 16

8

Question 17

Describe the TCA cycle.

Answer 17

A two carbon unit (from acetyl coA) condenses with a four carbon unit (oxaloacetate) to produce citrate (a 6 carbon unit). Citrate is oxidised and decarboxylated (which released a CO2 and produces an NADH + H+ from NAD+) 2 times to form a 4 carbon unit. Thus 2 CO2s have been released, and 2NADH + 2H+ have been produced.
A GTP ( which can be converted to ATP) is produced from GDP, by substrate level phosphorylation. There are 2 more oxidation reactions. In one of these oxidation reactions, FAD is reduced to FADH2, and in the other, NAD+ is reduced to NADH + H+.
Oxaloacetate is then regenerated and the cycle can start again.
In total, 3NADH + 3H+, 1 FADH2, one GTP and 2 CO2 are produced in one round of the TCA cycle.

Question 18

Which two molecules in the TCA cycle have 3 carboxyl groups? (I.e. are tricarboxylic)

Answer 18

Citrate and isocitrate

Question 19

What are the two ways of phosphorylating nucleotides?

Answer 19

Substrate level phosphorylation and oxidative phosphorylation.

Question 20

Where are the enzymes of the TCA cycle located?

What is the one exception, and what else makes this enzyme different?

Answer 20

They are all located in the matrix of the mitochondria, except for Succinate dehydrogenase, which is integrated in the inner mitochondrial membrane. This enzyme is also different because it is the one enzyme in the TCA cycle which uses FAD as a cofactor instead of NAD+.

Question 21

How do carbohydrates enter the TCA cycle?

Answer 21

They are converted to pyruvate, then Acetyl coA, and enter the TCA cycle in this form.

Question 22

How do lipids enter the TCA cycle?

Answer 22

Lipids are converted to fatty acids by beta oxidation. These are converted to acetyl coA, which enters the TCA cycle.

Question 23

How do proteins enter the TCA cycle?

Answer 23

Proteins are converted to amino acids. The amino acids are either converted to pyruvate, or to acetyl coA, or are converted to intermediates in the TCA cycle.

Question 24

How many substrate level phosphorylation reactions occur in the TCA cycle?

Answer 24

One, in the conversion of GDP to GTP.

Question 25

Which molecules inhibit the TCA cycle?
Remember- the TCA cycle has two functions: to provide energy, and to provide precursor molecules for biosynthetic reactions.

Answer 25

High ATP, NADPH and acetyl coA inhibits the TCA cycle, as these all mean that there is a large supply of energy.
High acetyl coA and high succinyl coA also inhibit the TCA cycle as these mean that there is a good supply of precursor molecules for biosynthetic reactions.

Question 26

Which molecules activate some steps in the TCA cycle?

Answer 26

High ADP and high NAD+ as these signify a lack of energy.

Question 27

What is generated in each turn of the TCA cycle?

Answer 27

3 x NADH + H+
1 x FADH2
1 x GTP
2 x CO2

Question 28

How can GTP be converted to ADP?

Answer 28

GTP + ADP –> GDP + ATP

Question 29

At the end of glycolysis and the TCA cycle, how many cofactors have now been reduced, and still have to be reoxidised (assuming we started with 1 glucose)?

Answer 29

10 x NADH + H+ (2 from glycolysis, 2 from conversion of pyruvate into acetyl coA [ this occurs twice for each glucose], 6 from the TCA cycle [which occurrs twice for each glucose].
2 x FADH2 (2 from the TCA cycle which occurs twice for each glucose)