W5 - Enzyme Kinetics

Question 1

What is a holoenzyme or an apoenzyme?

Answer 1

Holoenzyme - enzyme with co-factor, Apoenzyme - enzyme without co-factor

Question 2

Why does changing the activation energy not change the deltaG of the reaction?

Answer 2

The start and end energies remain the same

Question 3

Are all enzymes proteins?

Answer 3

No e.g. catalytic RNA

Question 4

How do enzymes lower the activation energy?

Answer 4

They stabilise the transition state

Question 5

What enzyme catalyses the conversion between carbon dioxide and carbonic acid?

Answer 5

Carbonic anhydrase

Question 6

Active sites are unique chemical microenvironments and are more likely hydrophobic, what 4 weak interaction occur at the AS?

Answer 6

Electrostatic, H-bonding, VdWs and hydrophobic

Question 7

Does making or breaking a bond release/require energy?

Answer 7

Making - releases, Breaking - requires

Question 8

Enzymes have no change on delta G and ???

Answer 8

Eq constant

Question 9

What is the equation for rate of reaction?

Answer 9

Change in products over change in time

Question 10

What is the units for rate?

Answer 10

Mol per sec (Ms-1) (Mol/L/s)

Question 11

What 6 things can determine RoR?

Answer 11

Conc, catalysts, temp, pressure, pH, number of collisions

Question 12

What is the rate equation for A + B –> P?

Answer 12

V = k [A][B]

Question 13

What are the units of k, the rate constant?

Answer 13

M-1 s-1

Question 14

Over time the RoR will go down, why?

Answer 14

The amount of substrate decreases as it’s converted to product

Question 15

At equilibrium, what is equal?

Answer 15

The rate of the forward and backwards reactions

Question 16

What is V0?

Answer 16

The initial (fastest) rate of reaction

Question 17

At a fixed enzyme conc, what happens to the RoR when the [substrate] increases?

Answer 17

RoR increases

Question 18

How to calculate V0?

Answer 18

Draw tangent on curve that goes through (0,0)

Question 19

What is on the x and y axis of the Michaelis Menten curve?

Answer 19

x - substrate conc, y - reaction velocity

Question 20

What graph/experiment do you need to do before making the M-M curve?

Answer 20

Conc of product against time at multiple different substrate concentration, calculate V0 for each (plotted on M-M curve)

Question 21

What is the Vmax on the M-M curve?

Answer 21

Where the graph plateaus

Question 22

What is the M-M formula?

Answer 22

V = Vmax ( [S] / ([S] + Km)) where Km is the Michaelis constant

Question 23

What are the 4 things that give evidence for ES complexes?

Answer 23

Structural data from crystallography, spectroscopic data, electrophoretic crossing line and saturation effect

Question 24

How does the electrophoretic crossling line experiment give evidence for the ES complex?

Answer 24

As the complex forms the molecule gets larger and doesn’t travel as far

Question 25

What are the 2 reversible reactions of enzyme?

Answer 25

E+S forms ES, ES forms E + P

Question 26

What are the rate constants for each reaction in enzyme action?

Answer 26

E+S --> ES has rate constant of K1 and the reverse rxn has K-1, ES --> E + P has rate constant of K2 and the reverse rxn has K-2

Question 27

Why can we ignore K-2 in the initial rxn?

Answer 27

The reverse rxn is minimal

Question 28

What is Vmax?

Answer 28

Theoretical maximum rate of reaction

Question 29

What is the equation for Vmax?

Answer 29

Vmax = K2[ET] where [ET] is total enzyme conc

Question 30

Describe what the M-M curve tells you?

Answer 30

The rate is initially proportional to [S] but it reaches a saturation value (Vmax)

Question 31

At Vmax the ES --> E+P part of enzyme action determines how quickly the enzyme catalyses the reaction, what is the K2 also known as?

Answer 31

The Kcat

Question 32

What do you call the enzymes turnover reaction?

Answer 32

Catalytic power

Question 33

What 3 things does Km depend on?

Answer 33

pH, temp and ionic strength

Question 34

Km is equal to the conc of substrate required for the reaction velocity to be ....

Answer 34

Half it's maximum value

Question 35

When [S] = Km, then V =

Answer 35

Vmax/2

Question 36

What does the Km tell you about the M-M curve?

Answer 36

The smaller the Km value the steeper the gradient of initial Ror

Question 37

What is the equation for Km?

Answer 37

Km = (K-1 + K2) / K1

Question 38

If K2 < K-1, Km is equal to the...

Answer 38

Dissociation constant of the ES complex (Kp)

Question 39

Km tells us the affinity of ES complex, If the Km is large/small, what does this tell us about the binding in the ES complex?

Answer 39

Large - weak binding, small - strong

Question 40

What is the diffusion limit?

Answer 40

The maximum value for K2/Km (the catalytic efficiency constant)

Question 41

What does the value of K2/Km have to be between to be diffusion limited?

Answer 41

10^8 and 10^9

Question 42

How to find Km on a graph?

Answer 42

Find Vmax, then half Vmax and draw across then down to find Km

Question 43

If glucokinase acts faster but has a lower affinity for substrate and hexokinase acts slower but has higher affinity for substrate what does this tell you?

Answer 43

Glucokinase works for storage better at higher glucose levels and hexokinase works in muscles better at lower lower levels

Question 44

What does the Lineweaver -Burk plot, plot?

Answer 44

1/V0 against 1/[S]

Question 45

What does the gradient, the y intercept and the x - intercept of the L-B plot tell you?

Answer 45

Gradient = Km/Vmax, y - intercept - 1/Vmax, x - intercept - (-1/Km)

Question 46

If 1/Vmax goes up what happens to the Vmax?

Answer 46

It decreases

Question 47

If Vmax gets lower and Km remains the same what does it tell us about the activity and affinity of the ES?

Answer 47

Activity is lower and affinity is the same

Question 48

What enzymes do not follow the M-M kinetics?

Answer 48

Allosteric enzymes (produces sigmoidal graph instead)