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2.1. In order to measure the enzyme activity and the initial rate of reaction, 5 mL of cellobiose
(100mumol/mL) and 44 mL of buffer solution were placed in a stirred vessel. The reaction was
initiated by adding 1 mL of enzyme (beta-glucosidase) solution which contained 0.1 mg of
protein per mL. At 1, 5, 10, 15, and 30 minutes, 0.1 mL of sample was removed from the
reaction mixture and its glucose content was measured. The results were as follows:
a. What is the activity of the ß-glucosidase in units/mL of enzyme solution and in umts/mg
protein? A unit is defined as the enzyme activity which can produce 1 mumol of product
per minute.
b. What is the initial rate of the reaction?
Solution:
y = 0.033x + 0.039
1.65 𝑚𝑢𝑚𝑜𝑙/𝑚𝑖𝑛
a. Activity of the ß-glucosidase (units/mL) = 1 𝑚𝐿 𝑜𝑓 𝑒𝑛𝑧𝑦𝑚𝑒
y = 0.033x + 0.0391
1 R² = 0.9973
Concentration (µmol/mL)
0.8
0.6
0.4
0.2
0
0 5 10 15 20 25 30 35
Time (min)
2.5. Eadie (1942) measured the initial reaction rate of hydrolysis of acetylcholme (substrate)
by dog serum (source of enzyme) and obtained the following data:
Solution:
A. Langmuir Plot
Langmuir Plot
Cs Cs/R 0.06
y = 3.3133x + 0.0191
0.0032 0.028829 0.05 R² = 0.8837
0.0049 0.033108 0.04
Cs/R
0.0062 0.043357 0.03
y = 3.313305715x + 0.01912464593
1
m= = 3.313305715
rmax
Thus,
1
rmax =
3.313305715
= 0.301813381 mol L • min
KM
b= = 0.01912464593
rmax
Thus,
K M = b × rmax
= (0.01912464593)(0.301813381)
= 5.772074049 × 10 -3 mol L
Tangonan, Visitacion D. February 27, 2019
Biochemical Engineering
B. Lineweaver-Burk Plot
1/V
105.2632 5 4
0
0 50 100 150 200 250 300 350
I/Cs
1 1 K 1
m
V Vmax Vmax C s
y = 0.01719105838x + 3.63421308
1
b= = 3.63421308
rmax
Thus,
1
rmax =
3.63421308
= 0.2751627321 mol L • min
KM
m= = 0.01719105838
rmax
Thus,
K M = m × rmax
= (0.01719105838)(0.2751627321)
= 4.730338591 × 10 -3 mol
L
Tangonan, Visitacion D. February 27, 2019
Biochemical Engineering
C. Eadie-Hofstee Plot
Eadie-Hosftee Plot
R/Cs R 0.25
34.6875 0.111
0.2
30.20408 0.148
23.06452 0.143 R
0.15
20.75 0.166 0.1 y = -0.0043x + 0.2645
21.05263 0.2 R² = 0.6584
0.05
0
0 10 20 30 40
R/Cs
r
r = -K M + rmax
Cs
y= m x +b
y = -4.273141382 × 10 -3 x + 0.2644954792
b = rmax = 0.2644954792 mol L • min
m = -K M = 4.273141382 × 10 -3 mol L