Enzyme Kinetics Examples and Problems

1.

An enzyme is produced for producing a sun protection lotion. Given kinetic data for the enzyme reaction with Vm=2.5
mmol/m3.s, Km=8.9 mM and So=12mM, what would be the time required for 95% conversion in a batch reactor?

2.

An enzyme was assayed at an initial substrate concentration of 10-5M. The Km for the substrate is 2x10-3M. At the end of 1
min, 2% of the substrate had been converted to product.
a. What % of the substrate will be converted to product at the end of 3 min.? calculate the product and substrate
concentration at that time.
b. If the initial concentration of the substrate were 10-6M, what percent of the substrate will be converted to product
after 3 min?
c. What is the maximum attainable velocity of the reaction with the enzyme concentration used?
d. At about what substrate concentration will Vm be observed?
e. At this S concentration, what % of the substrate will be converted to product after 3 min?

3.

An enzyme catalyzed reaction of the form S P has a value of G0 = 3.4 kJ/mole.

a. Calculate the equilibrium constant at 298K
b. If we start with a solution containing 0.1M substrate, what is the at equilibrium?

4.

Given the reaction E + S - E.S -- E + P where k1 = 1x107M/s, k-1=1x102 /s and k2=3x102/s

a. Calculate Km, Km

5.

An enzymatic assay was carried under two different sets of conditions out using a pure substrate S. The results are tabulated
below.
a. Calculate the Km and Vm for given substrate concentrations and rates for A only.
b. Use the 3 plots and comment on their differences
c. For inhibition example do B then compare with A
A

S (mol/L)

V0 (mol/L.min)

v (mol/L.min)

1.5x10-5

0.21

0.08

2.0x10

-5

0.25

0.1

3.0x10

-5

0.28

0.12

4.0x10

-5

0.33

0.13

8.0x10-5

0.44

0.16

0.40

0.18

16.0x10
6.

-5

You are given a 4.0 ug/mL solution of an enzyme (MW = 40,000Da). In your experiment, you have determined the initial
reaction velocities vo at several substrate concentrations, [S].
a. Determine the Km and Vm using lineweaver-Burke Plot
b. Calculate the turnover number for the reaction
S (mM)

V0 (mM/min)

0.25

0.06

0.4

0.08

0.7

0.13

0.16

0.25

0.31

0.36

0.39

12
7.

8.

w/o

with

S (mol/L)

v (mol/L.min)

v (mol/L.min)

3.30E-04

5.60E-05

3.70E-05

5.00E-04

7.10E-05

4.70E-05

6.70E-04

8.80E-05

6.10E-05

1.65E-03

1.29E-04

1.03E-04

2.21E-03

1.49E-04

1.25E-04

Rate measurements were made on a food enzyme with and without presence of an inhibitor with the following results.
Inhibitor concentration is [I] = 20uM

a.
b.
c.
9.

0.43

20
0.46
Pesticide inhibition on an active enzyme has been reported which caused enzyme activities to reduce. Collected data are
presented for both without and with inhibition.
a. Determine the rate model with and without inhibitor.
b. Determine the type of inhibition.

w/o

with

S/uM

v, umol/min

v, umol/min

25

8.2

5.65

50

12.86

8.16

75

15.51

9.68

100

17.95

11.04

150

20.02

13.56

200

22.64

15.71

Use either lineweaver-Burk or Eadie-Hoftstee plot to determine type of inhibition

Determine Km and Vo for the enzyme without inhibition as well as K3, the inhibitor constant
What further tests are needed to improve the accuracy of the estimate of K3? How would you calculate then?

An experiment on the thermal degradation of an enzyme was carried out and some of the results are below

a.
b.
c.

Time, s

%Activity @ 65C

%Activity @75C

100

100

100

92

85

200

85

72

500

67

45

1000

45

20

Plot a graph of log activity vs. time and determine the inactivation rate constants at 65C and 75C.
Determine the activation energy for the thermal degradation of the enzyme.
When eggs are pasteurized, alpha-amylase activity is often used as an indicator that the pasteurization process has been
satisfactorily completed. Suggest a reason why alpha-amylase activity may be used in this way.

10. The equilibrium constant for the reaction S- P is 5. Suppose we have a mixture of [S] = 2x10-4M and [P] = 3x10-4M. Km
= 3x10-5M, Vm=2 umol/(L.min), Vm=4 umol/(L.min)
a. Which direction will the reaction proceed on addition of an appropriate enzyme?
b. At what initial velocity will the reaction start towards equilibrium?
11. Estimate the k, the first order rate constant, for an enzyme preparation with a Vm = 4.6 umol/L.min under the given
experimental condition, Km = 2x10-6M

12. An enzyme was assayed at an initial substrate concentration of 2x10-5M. In 6 min, half of the substrate had been used. The
Km for substrate is 5x10-5M.
a. Calculate k, Vm, and concentration of product by 15 min.
13. An enzyme has a Km of 4.7x10-5M. If the Vm of the preparation is 22 umol/L.min, what velocity would be observed in the
presence of 2x10-4M substrate and 5x10-4M of;
a. Competitive inhibitor
b. Non competitive inhibitor
c. Uncompetitive inhibitor
If Ki in all cases is 3x10-4M
14. The effect of temperature on the hydrolysis of lactose by a beta-galactose is shown below
a. Calculate the activation energy
T, 0C
Vm (umol/mg protein.min)
20

4.5

30

8.65

35

11.8

40

15.96

45

21.36