Lecture 2

Question 1

What are enzymes an what are their general purposes?

Answer 1

They are proteins that are biological catalysts. They control metabolic reactions, each reaction has a highly specific enzyme to do the job. Some enzymes however require metal ions or organic cofactors to become active.

Question 2

What are the 6 main classifications of enzyme. And what type of reaction do they catalyse?

Answer 2

Oxidoreductases - Oxidation-reduction reactions, Transferases - Transfer of functional groups, Hydrolases - Hydrolysis reactions, Lyases - Group elimination to form double bonds, Isomerases - Isomerisation and Ligases - Bond formation and ATP hydrolysis

Question 3

What does the complementarity of the enzyme and the substrate depend on?

Answer 3

Non-covalent forces

Question 4

What are the basic steps to how an enzyme works?

Answer 4

1. The substrate enters the enzymes active site
2. The enzyme-substrate complex (ESC) is formed
3. The enzyme-product complex is formed
4. The products of the reaction leave the active site of the enzyme

Question 5

How does the enzyme speed up the rate of a reaction?

Answer 5

It lowers the activation energy needed for the reaction by providing a lower energy pathway from substrate to product

Question 6

In an enzyme catalysed reaction, what stays unchanged?

Answer 6

There is no overall effect on the free energy change of the reaction

Question 7

What does studying enzyme kinetics help us to understand?

Answer 7

• Helps us to understand the mechanisms of enzymes
• Helps us to understand the regulation of biochemical reactions in metabolic pathways
• Helps us to investigate mutations of the metabolic pathways

Question 8

How does the concentration of substrate affect the initial rate of reaction when the enzyme concentration is constant and low?

Answer 8

Low concentration - The rate of the reaction is dependent on the substrate concentration, so the binding of the substrate is the limiting factor
High concentration - The rate of the reaction is independent of the substrate concentration, the enzymes are fully saturated

Question 9

What is the Michaelis-Menten kinetic equation?

Answer 9

Vo = (Vmax x [S])/ (Km + [S])

Question 10

What is the Michaelis constant [Km] and what does it show?

Answer 10

It shows the substrate concentration at which the reaction reaches half of its maximum velocity (Vmax/2)

It also measures the affinity of the enzyme for the substrate

Question 11

What does Vmax of an enzyme catalysed reaction show?

Answer 11

Shows the initial reaction velocity when the substrate concentration is high compared to the Km

Question 12

What happened to the Michaelis-Menten equation?

Answer 12

In the 1930s Burk and Lineweaver found limitations to the equation and overcame the limitations of Vmax determination by linearising the equation

Question 13

What was the Lineweaver-Burk equation and what type of graph did they plot to determine the values needed?

Answer 13

1/Vo = (Km/Vmax) x (1/[S] + 1/Vmax)

They created a ‘double reciprocal’ plot

Question 14

How are enzymes controlled within an organism?

Answer 14

Through the enzymes availability - Controlling the rate of synthesis and degradation

Through the enzymes activity - Inhibiting or modifying the enzyme

Question 15

What is the effect of pH on enzyme activity?

Answer 15

• Higher activity at a narrow pH range (optimum pH)
• The R groups of the Amino Acids that make up the enzyme can get changed at different pH levels
• In too low or too high of a pH the tertiary structure of the enzyme is disrupted and the enzymes lose their activity (denatured)

Question 16

What is the effect of temperature on enzymes?

Answer 16

A range of temperatures are considered optimal for enzymes, most have an optimum temperature of around 37 degrees and if the temperature is too high the enzyme will degrade and become denatured

Question 17

Why is the inhibition of enzymes studied?

Answer 17

• Many therapeutic drugs are inhibitors of specific enzymes

- We can use inhibition as a probe for understanding the kinetic and properties of enzymes in their uninhibited state

Question 18

What is the definition of an enzyme inhibitor?

Answer 18

A substance that reduces an enzymes activity by influencing the binding of substrate and/ or its reaction rate

Question 19

What does the pharmaceutical industry refer to enzyme inhibitors as?

Answer 19

Novel drugs

Question 20

What are the 3 types of enzyme inhibitors?

Answer 20

• Competitive inhibitors
• Non-competitive inhibitors
• Uncompetitive inhibitors

Question 21

What is the mechanism of action for a competitive inhibitor?

Answer 21

The inhibitor competes directly with the substrate for the same enzyme binding site

Question 22

What is the mechanism of action for a non-competitive enzyme. Inhibitor?

Answer 22

Inhibitor binds to the enzyme allosteric site (not the active site) and prevents the binding of the substrate

Question 23

What is the mechanism of action of an uncompetitive inhibitor on enzymes?

Answer 23

The inhibitor binds to the enzyme however the substrate can still bind to the active site, although nothing will happen as the inhibitor inactivates the enzyme completely

Question 24

What happens to the Km and the Vmax values of an enzyme catalysed reaction when there is a competitive inhibitor present?

Answer 24

The Km is increased but the Vmax is unaffected

Question 25

What happens to the Km and the Vmax values of an enzyme catalysed reaction when there is an uncompetitive inhibitor present?

Answer 25

The Km and the Vmax values both decrease

Question 26

What happens to the Km and the Vmax values of an enzyme catalysed reaction when there is a non-competitive inhibitor present?

Answer 26

The Km value stays unaffected, however the Vmax value decreases

Question 27

Why are inhibitors used as drugs?

Answer 27

• Some inhibitors can be used to stop enzymes that are necessary for the functioning of a pathogen
• Some can be used for enzymes that are unregulated in a human disease (Disease biomarkers)
• Some can be used to target enzymes that are produced more energetically by tumours than by normal tissues (Anti cancer drugs)

Question 28

What is similar between the competitive inhibitors and the substrate?

Answer 28

The chemical structure has to be very similar as it needs to be able to bind to the same active site of the enzyme

Question 29

Enzymes can also be regulated by covalent phosphorylation, how does this happen?

Answer 29

It happens at the -OH group (Ser,Thr and Tyr residues)
Involves using ATP and a Kinase
The inverse process (dephosphorylation) is catalysed by phosphatases

Question 30

Give an example of an enzyme that is found in the blood and used to diagnose human disease?

Answer 30

PSA (Prostate-specific antigen) high levels in the blood can be a sign of prostate cancer