Patrick McCrystal

Enzymes: Natural Catalysts

Enzymes are catalytic proteins, meaning they speed up chemical reactions without being
used up or altered permanently in the process. Although various enzymes use different methods,
all accomplish catalysis by lowering the activation energy for the reaction, thus allowing it to
occur more easily. Enzymes have very specific shapes (conformations). Part of the conformation
is the active site of the enzyme, where the actual catalysis occurs. The specific molecule or
closely related molecules on which an enzyme functions is known as its substrate. Shape plays
such an important role in enzymatic catalysis that often even isomers of a substrate will be
rejected. Once the substrate enters the active site, it may begin a process known as induced fit in
which the enzyme perfectly conforms to the molecule to allow for more efficient catalysis.
Changes in environment can severely impact enzyme catalysis in both negative and positive
ways. Each enzyme has specific ranges at which it optimally functions; in general, increasing the
temperature will help the reaction along, until the point at which the protein degrades and
denatures. Denatured proteins will often return to their original state, after the removal of the
denaturing agent, except when they are degraded multiple levels.

Methods:
1. Peel a fresh potato tuber and cut the tissue into small cubes.
2. Weigh out 50 grams of tissue.
3. Place the tissue, 50 mL of cold distilled water, and a small amount of crushed ice in a
prechilled blender.
4. Homogenize for 30 seconds at high speed.
5. Filter the potato extract using cheesecloth.
6. Pour the filtrate into a 100 mL graduated bylinder and add cold distilled water to bring up
the final volume to 100 mL.
7. Label eight 50 mL beakers as follows: 100 units/mL, 80 units/mL, 75 units/mL, 60
units/mL, 50 units/mL, 25 units/mL, 10 units/mL, 0 units/mL.
8. Prepare 40 mL of enzyme for each of the above concentrations in the following ratio of
enzyme:distilled water – 40:0, 32:8, 30:10, 24:16, 20:10, 10:30, 4:36, and 0:40.
9. Using forceps, immerse a 2.1cm filter paper disc into the prepared catalase solution for 5
seconds.
10. Remove the disc and drain for 10 seconds on a paper towel.
11. Place the disc at the bottom of the first substrate solution. The oxygen produced from the
breakdown of the hydrogen peroxide by catalase becomes trapped in the fibers of the
disc, thereby causing the disc to float to the surface of the solution
12. Measure (using a stopwatch) the reaction time for the amount of time from when the disc
was placed at the bottom of the beaker until the disc floats on top of the solution. (rate
will be measured in seconds). The rate (R) of the reaction is calculated as R = 1/t.
13. Repeat this procedure twice for each enzyme concentration and average the results.
Data:
Enzyme Time to Float Disc
Concentration (seconds)
(units/mL) Trial 1 Trial 2 Average Rate
100 3.4 3.07 3.235 1/3
80 2.69 2.66 2.675 1/3
75 2.54 2.39 2.465 1/3
60 5.85 6.21 6.03 1/6
50 6.17 4.48 5.325 1/5
25 11.56 15.69 13.625 1/14
10 17.83 30.17 24 1/24
0 ∞ ∞ ∞ 1/∞

Effect of Enzyme Concentration on

Rate of Activity

35
Time to Float Disc (sec)

30
25
Trial 1
20
Trial 2
15
Average
10
5
0
1 2 3 4 5 6 7
Enzyme Concentration (units/mL)

Substrate Time to Float Disc

Concentration (seconds)
(from serial dilution) Trial 1 Trial 2 Average Rate
0% ∞ ∞ ∞ 1/∞
0.10% 192.07 160.37 176.22 1/176
0.20% 47.89 52.05 49.97 1/50
0.30% 48.29 48.81 48.55 1/49
0.50% 31.92 37.91 34.915 1/35
0.80% 20.61 26.55 23.58 1/24
1% 14.64 15.8 15.2 1/15
2% 2.59 8.13 5.36 1/5
3% 5.8 5.08 5.44 1/5
 Effect of Substrate Concentration on
Enzyme Activity

250
Time to Float Disc

200
(seconds)

Trial 1
150
Trial 2
100
Average
50
0

1%

2%

3%
%

%
%

%
10

20

30
50

80
0.

0.

0.
0.

0.

Substrate Concentration (from

serial dilutions)

Conclusion:
This lab was helpful in showing us how enzyme catalysis really happens in biology. To
get better results, we should have done this lab over again, but time did not allow us to do so.
The results we got, however, were roughly to be expected. The experiments showed a trend of
lower times to float the disc as you increased the concentration of enzyme/substrate. Errors could
have occurred in many ways. The potato enzyme may have been improperly diluted before its
addition to distilled water. Students may have measured their dilutions incorrectly. The enzyme
may have not been kept cold enough until its use. Students may have improperly recorded the
time in which it took for the disc to float. Enzymatic processes are necessary to life, as can be
seen to their role in digestion, among other things. This lab taught students about the intricacies
of enzymatic activity and its importance in everyday life.