S4) Enzymes

Question 1

What are enzymes and what do they do?

Answer 1

Enzymes are protein catalysts that mediate metabolic pathways by increasing the velocity of chemical reactions while not being consumed during the reaction

Question 2

Identicy 5 characteristics of enzymes

Answer 2

• Highly specific
• Unchanged after reaction
• Increase rate of reaction
• Proteins
• Does not affect reaction equilibrium

Question 3

What is an active site?

Answer 3

An active site is the special cleft on an enzyme containing amino acid side chains that bind substrates and trigger catalysis

Question 4

Identify 4 characteristics of active sites

Answer 4

• Cccupies a small part of the enzyme
• Formed by amino acids
• Clefts/crevices
• Bind to substrates through weak bonds

Question 5

Identify 2 different hypotheses which govern substrates binding to an enzyme

Answer 5

• Lock and key hypothesis – active sites have a complementary shape to the substrate
• Induced fit hypothesis – binding of substrates can induce chages in the comformation of the active site

Question 6

Describe the 5 steps involved in enzyme catalysis

Answer 6

• Substrate binds to active site on enzyme
• Conformational change occurs and ES complex forms
• Catalysis occurs
• ES complex is converted to EP complex
• EP compex dissociates to enzyme and product

Question 7

What is an allosteric site?

Answer 7

An allosteric site is a cleft on an enzyme where an activating/inhibiting molecule may bind, changing the shape of the enzyme and influencing its ability to be active

Question 8

Outline the specificity of enzymes

Answer 8

• Enzymes are highly specific, interacting with 1/few substrates and catalysing only one type of chemical reaction
• The set of enzymes made in a cell determines which metabolic pathways occur in that cell

Question 9

Can enzymes be regulated?

Why is this needed?

Answer 9

Enzyme activity can be regulated (increased or decreased) so that the rate of product formation responds to cellular need

Question 10

Why are many enzymes localised in specific organelles within the cell?

Answer 10

Compartmentalisation serves to:

• Isolate the reaction substrate/product from other competing reactions
• Provides a favourable environment for the reaction
• Organises thousands of cell enzymes into purposeful pathways

Question 11

What is activation energy?

Answer 11

Activation energy is the minimum energy required by a chemical system to start a chemical reaction

Question 12

How do enzymes impact activation energy in a chemical reaction?

Answer 12

• Activation energy is usually very high, leading to slow rates in uncatalyzed chemical reactions
• Enzymes work to lower activation energy, so more particles are able to overcome the transition state and react
• Rate of reaction increases

Question 13

What is the rate of reaction?

Answer 13

The rate of a reaction is the number of substrate molecules converted to product per unit time ( [product]/time )

Question 14

How does the rate of reaction vary in the presence of an enzyme?

Answer 14

The rate of an enzyme-catalysed reaction increases with substrate concentration until a maximal velocity (Vmax) is reached

Question 15

Why does the rate of reaction level off in enzyme-catalysed reactions?

Answer 15

The levelling off of the reaction rate at high substrate concentrations reflects the saturation of all available enzyme active sites with substrate

Question 16

Identify and describe 2 factors which increases the rate of reaction

Answer 16

• Temperature: increases no. of molecules with activation energy
• Concentration: increases chance of molecular collisions

Question 17

In Michaelis-Menten kinetics, what type of curve is shown when measuring the rate of reaction for an enzyme-catalysed reaction?

Answer 17

Rate of reaction is hyperbolic

Question 18

For allosteric enzymes, what type of curve is shown when measuring the rate of reaction for an enzyme-catalysed reaction?

Answer 18

Rate of reaction is sigmoidal e.g. oxygen dissociation curve of haemoglobin

Question 19

Discuss the effect of temperature on reaction velocity

Answer 19

• Initially, an reaction velocity increases with temperature due more molecules surpassing the energy barrier of the reaction
• Reaction velocity then decreases due to the temperature-induced denaturation of the enzyme

Question 20

What is the optimum temperature for most human enzymes?

Answer 20

Between 35 and 40°C

Question 21

Discuss the effect of pH on enzyme function

Answer 21

Extremes of pH lead to enzyme denaturation due to a conformational change the structure of enzyme (active site)

Question 22

What is the optimum pH for human enzymes?

Answer 22

The optimum pH varies for different enzymes and often reflects the [H⁺] at which the enzyme functions in the body

Question 23

State the Michaelis-Menten equation

Answer 23

• V = initial reaction velocity
• V_max = maximal velocity
• K_m = Michaelis constant
• [S] = substrate concentration

Question 24

What is K_m in Michaelis-Menten kinetics?

Answer 24

• K_m is numerically equal to the substrate concentration at 1⁄2 V_max
• Km does not vary with the concentration of enzyme

Question 25

What is the significance of K_m?

Answer 25

- K_m is characteristic of an enzyme and its particular substrate - It reflects the affinity of the enzyme for that substrate

Question 26

Compare and contrast the enzyme affinity in terms of a small K_m and large K_m

Answer 26

- **Small K_m:** high affinity of the enzyme for a substrate as a low [substrate] is needed to half-saturate the enzyme - **Large Km:** low affinity of enzyme for substrate as a high [substrate] is needed to half-saturate the enzyme

Question 27

Describe the relationship of velocity to enzyme concentration in Michaelis-Menten kinetics

Answer 27

The rate of the reaction is **directly proportional** to the enzyme concentration at all substrate concentrations ## Footnote *E.g. if the [enzyme] is halved, the rate of reaction is halved*

Question 28

What is a Lineweaver-Burk plot?

Answer 28

A **Lineweaver-Burk plot** plots 1/V0 versus 1/[S] to obtain a straight line graph for Michaelis-Menten kinetics - It is used to calculate K_m and V_max, as well as to determine the mechanism of action of enzyme inhibitors

Question 29

What are the values of the x and y intercepts in a Lineweaver Burk plot?

Answer 29

- **X-intercept** is equal to −1/Km **- Y-intercept** is equal to 1/Vmax

Question 30

What is an inhibitor?

Answer 30

An **inhibitor** is any substance that can diminish the velocity of an enzyme-catalysed reaction

Question 31

Distinguish between the bonding of irreversible and reversible inhibitors

Answer 31

- Irreversible inhibitors bind to enzymes through **covalent bonds** - Reversible inhibitors bind to enzymes through **non-covalent bonds**

Question 32

Identify 2 forms of reversible inhibition

Answer 32

- Competitive inhibition - Non-competitive inhibition

Question 33

Outline competitive inhibition

Answer 33

- Inhibitor binds reversibly to the same site the substrate would occupy - Inhibitor competes with the substrate for the active site - Increasing [substrate] reduces the effect of inhibitor

Question 34

Describe the effect of competitive inhibition on the V_max and K_m

Answer 34

- **Effect on V_max:** effect is reversed by increasing [S] & rate of reaction reaches V_max - **Effect on K_m:** increases the K_m for a given substrate as more substrate is needed to achieve 1⁄2 V_max

Question 35

Describe the Lineweaver-Burk plot for competitive inhibition

Answer 35

- Inhibited and uninhibited reactions **intersect on the y-axis** at 1/V_max (V_max is unchanged) - The inhibited and uninhibited reactions show **different x-axis intercepts** (K_m is increased)

Question 36

Outline non-competitive inhibition

Answer 36

- Inhibitor binds reversibly to allosteric site on the enzyme - Inhibitor does not compete with the substrate for the active site - Increasing [substrate] has no effect on inhibition as active site has changed shape

Question 37

Describe the effect of competitive inhibition on the V_max and K_m

Answer 37

- **Effect on V_max:** decrease the V_max of the reaction as inhibition is not overcome with increasing [substrate] - **Effect on K_m:** same value of K_m as inhibitors do not interfere with the binding of substrate to enzyme

Question 38

Describe the Lineweaver-Burk plot for non-competitive inhibition

Answer 38

- Inhibited and uninhibited reactions **intersect on the x-axis** at 1/K_m (K_m is unchanged) - The inhibited and uninhibited reactions show **different y-axis intercepts** (V_max is decreased)

Question 39

What are amyloid fibres?

Answer 39

An **amyloid fibre** is the misfolded, insoluble form of a normally soluble protein

Question 40

Describe the structure of amyloid fibres

Answer 40

- Highly ordered with a high degree of ß-sheet - Core ß-sheet forms before the rest of the protein - Stabilised by hydrophobic interactions between aromatic amino acids