BIOLOGY 6
ENZYME LAB
NAME _____________________________
At any given moment, all of the work being done inside any cell is being done by enzymes. If
you understand enzymes, you understand cells. A bacterium like E. coli has about 1,000 different
types of enzymes floating around in the cytoplasm at any given time. A living system controls its
activity through enzymes. An enzyme is a protein molecule that is a biological catalyst with
three characteristics. First, the basic function of an enzyme is to increase the rate of a reaction.
Most cellular reactions occur about a million times faster in the presence of an enzyme. Second,
most enzymes act specifically with only one reactant (called a substrate) to produce products.
The third and most remarkable characteristic is that enzymes are regulated from a state of low
activity to high activity and vice versa. Gradually, you will appreciate that the individuality of a
living cell is due in large part to the unique set of some 3,000 enzymes that it is genetically
programmed to produce. If even one enzyme is missing or defective, the results can be
disastrous.
Enzyme activity can be affected by other molecules. Inhibitors are molecules that decrease
enzyme activity; activators are molecules that increase activity. Many drugs and poisons are
enzyme inhibitors. Activity is also affected by temperature, pressure, chemical environment
(e.g., pH), and the concentration of substrate. Some enzymes are used commercially, for
example, in the synthesis of antibiotics. In addition, some household products use enzymes to
speed up biochemical reactions (e.g., enzymes in biological washing powders break down
protein or fat stains on clothes; enzymes in meat tenderizers break down proteins into smaller
molecules, making the meat easier to chew).
Enzymes are made from amino acids, and they are proteins. When an enzyme is formed, it is
made by stringing together between 100 and 1,000 amino acids in a very specific and unique
order. The chain of amino acids then folds into a unique shape. That shape allows the enzyme to
carry out specific chemical reactions -- an enzyme acts as a very efficient catalyst for a specific
chemical reaction. The enzyme speeds that reaction up tremendously.
For example, the sugar maltose is made from two glucose molecules bonded together. The
enzyme maltase is shaped in such a way that it can break the bond and free the two glucose
pieces. The only thing maltase can do is break maltose molecules, but it can do that very rapidly
and efficiently. Other types of enzymes can put atoms and molecules together. Breaking
molecules apart and putting molecules together is what enzymes do, and there is a specific
enzyme for each chemical reaction needed to make the cell work properly.
Maltose is made of two glucose molecules bonded together (1). The maltase enzyme is a protein that
is perfectly shaped to accept a maltose molecule and break the bond (2). The two glucose molecules
are released (3). A single maltase enzyme can break in excess of 1,000 maltose bonds per second,
and will only accept maltose molecules.
You can see in the diagram above the basic action of an enzyme. A maltose molecule floats near
and is captured at a specific site on the maltase enzyme. The active site on the enzyme breaks
the bond, and then the two glucose molecules float away.
You may have heard of people who are lactose intolerant, or you may suffer from this problem
yourself. The problem arises because the sugar in milk -- lactose -- does not get broken into its
glucose components. Therefore, it cannot be digested. The intestinal cells of lactose-intolerant
people do not produce lactase, the enzyme needed to break down lactose. This problem shows
how the lack of just one enzyme in the human body can lead to problems. A person who is
lactose intolerant can swallow a drop of lactase prior to drinking milk and the problem is solved.
Many enzyme deficiencies are not nearly so easy to fix.
Inside a bacterium there are about 1,000 types of enzymes (lactase being one of them). All of the
enzymes float freely in the cytoplasm waiting for the chemical they recognize to float by. There
are hundreds or millions of copies of each different type of enzyme, depending on how important
a reaction is to a cell and how often the reaction is needed. These enzymes do everything from
breaking glucose down for energy to building cell walls, constructing new enzymes and allowing
the cell to reproduce. Enzymes do all of the work inside cells.
Lab Activity
Activity 1: Observation of catalase activity on hydrogen peroxide (H2O2)
Direction: Place 1 mL of hydrogen peroxide into a clean plastic weighing tray and add
0.5 mL of catalase to it. Record what you observe.
Activity 2: Observation of hydrogen peroxide on potato slice AND banana.
Direction: Add 1 mL of hydrogen peroxide onto a slice of potato and banana and
describe what you observe.
Activity 3: Observation of hydrogen peroxide on skin.
Direction: Add 0.5 mL of hydrogen peroxide onto your skin and describe what you
observe. If you have a fresh cut, it’s best to put the peroxide on the cut.
Activity 4: Effects of Temperature On Catalase Activity
Direction:
Obtain 5 test tubes and label the tubes according to the table below. Pipet into each tube
exactly 4 ml of the catalase solution. Place the tubes in their respective temperature
condition for 5 minutes and remove immediately after 5 minutes. Next add 6 ml of
hydrogen peroxide to tube number 1 and determine catalase activity by measuring the
amount of oxygen that is produce by the enzymatic activity. Repeat with tubes 2 to 5.
Tube
Temperature (Celsius)
1
0
2
5
3
20
4
40
5
80
Amount of O2 (ml)
MEASURING OXYGEN PRODUCTION
PROCEDURES:
These steps have to be carried out quickly to prevent the oxygen
produced from escaping.
1. For tube 1, use a 10mL transfer pipet and pipet pump to draw up
10mL of the mixture.
2. Cover the tip of the 10mL pipet with your finger and remove the
pipet pump.
3. Cover the top of the 10mL pipet with a pipet bulb or stopper.
4. Place the filled and sealed 10mL pipet into the labeled test tube and
place the tube into a test tube rack.
5. Repeat steps 1-4 for the other tubes.
6. Allow for oxygen to collect in the pipet for 1 hour. Measure gas
production as instructed by the professor. Record your results.
7. When you are finished, clean your work area and dispose of the
contents of each tube as instructed by the professor.
Activity 5: Effects of pH On Catalase Activity
Direction:
Obtain 5 test tubes and label the tubes according to the table below. Pipet into each tube
exactly 4 ml of the catalase solution. Next pipet exactly 1 ml of the pH solutions into the
respective tubes and let sit for 5 minutes. After 5 minutes, add 5 ml of hydrogen peroxide
to tube number 1 and determine catalase activity by measuring the amount of oxygen that
is produced by the enzymatic activity. Repeat with tubes 2 to 5. Follow procedures from
Activity 4 to measure oxygen production.
Tube
pH
1
2
2
4
3
6
4
8
5
10
Amount of O2 (ml)
Activity 6: Effects of Temperature on Amylase Activity
Direction:
Obtain 5 test tubes and label the tubes according to the table below. Pipet into each tube
exactly 1 ml of the amylase solution. Place the tubes in their respective temperature
condition for 5 minutes and remove immediately after 5 minutes. Next add 2 ml of starch
solution to all tubes and let sit for one hour. After one hour, add 1 drop of Lugol’s iodine
and mix well. Record the color intensity of the tubes from 0 to 10. Repeat with tubes 2-5.
If amylase did not digest starch, the color of the tubes would be bluish black after the
addition of Lugol’s iodine. If amylase digested starch, the color of the tubes would be
less dark.
Tube
Temperature (Celsius)
1
0
2
5
3
20
4
40
5
80
% Transmission or Color Intensity
(0-10)
Activity 7: Effects of pH on Amylase Activity
Direction:
Obtain 5 test tubes and label the tubes according to the table below. Pipet into each tube
exactly 1 ml of the amylase solution. Add 0.5 ml of the respective pH solutions to each
of tube and let sit for 5 minutes. After 5 minutes, add 2 ml of starch solution to all tubes
and let sit for one hour. After one hour, add 1 drop of Lugol’s iodine and mix well.
Record the color intensity of the tubes from 0 to 10. Repeat with tubes 2-4. If amylase did
not digest starch, the color of the tubes would be bluish black after the addition of
Lugol’s iodine. If amylase digested starch, the color of the tubes would be less dark.
Tube
pH
1
2
2
4
3
6
4
8
5
10
% Transmission or Color Intensity (010)
Questions
_____1. An enzyme is what type of biological molecule?
a. Protein
b. Carbohydrate
c. Lipid
d. Starch
_____2. Amlyase is an example of a(n)
a. Carbohydrate
b. Simple sugar
c. Enzyme
d. Indicator
_____3. The substrate upon which catalase acts is
a. Starch
b. Liver
c. Water
d. Hydrogen Peroxide
_____4. The enzyme that breaks down hydrogen peroxide and is present in all cells is
a. Amylase
b. Catalase
c. Pepsin
d. Water
_____5. In the Enzyme lab, which of the following occurred?
a. Starch and sugar were both detected in the first part.
b. Only glucose was detected in the first part.
c. Glucose was only detected after the amylase was added.
d. The glucose was broken down into starch molecules.
_____6. At what pH does catalase enzyme work best?
a. Neutral
b. Basic
c. Acidic
d. Complex
_____7. Pepsin is an enzyme that occurs in the stomach. Upon which substance does it act?
a. Carbohydrate
b. Sugar
c. Glucose
d. Protein
_____8. When considering enzyme action vs. surface area, in which of the following would the enzyme
be most efficient?
a. Whole potato
b. Diced potato
c. Mashed potato
d. French fry
_____9. Which of the following temperatures would show a decrease in efficiency in enzyme action?
a. 25 degrees Celsius
b. 120 degrees Celsius
c. – 20 degrees Celsius
d. a & b
e. b & c
_____10. An enzyme works on how many different substrates?
a. 1
b. 2
c. 3
d. 4
_______11. Amylase is an enzyme that allows the human body to digest starch. Which of these diagrams
best represents part of the structure of amylase?
A.
B.
C.
D.
_______12. A dog gets many nutrients from its food including amino acids. Which of these can be built
_________13. Cells in the stomach produce pepsin, an enzyme, to help digest food. Pepsin works best at
a pH of 2. Which of these graphs most likely shows what will happen to the activity of pepsin as the pH
of the stomach is increased?
A
B
C
D
________14.
If the temperature in this experiment were reduced by ten degrees, what would most likely happen to
the rate of reaction?
A.
B.
C.
D.
It would increase.
It would decrease.
It would remain the same.
It would increase, then decrease
15. The table below lists enzymes that function in different locations in the human body, and the normal
pH and temperature ranges of these locations.
Use your understanding of the structure and function of enzymes to:
a) Describe how the activity of pepsin will change after it moves from the stomach to the small
intestine
b) Describe how changes in pH and temperature affect enzyme activity
c) Predict how a fever of 40°C would affect enzyme activity
16.
Label the following diagram:
happening in each diagram:
Describe what is