(Modified from A. Goodwin)
Name: ______________________________
Concepts in Biology (BIOL 100)
Spring 2013
02. Enzymes and Lactose Intolerance
Learning Objectives:
• To understand how substrate specificity is determined by an enzyme’s 3D structure.
• To understand how temperature, pH, substrate concentration and enzyme concentration
affect reaction effectiveness, and be able to explain why.
• To be able to discuss lactose intolerance in the context of enzyme activity.
Enzyme Basics
Most chemical reactions in a cell are mediated by proteins called enzymes. An enzyme has a
specific three-dimensional shape that determines which molecule/molecules, or substrates, it will
recognize. When an enzyme mediates molecule-building (anabolism), two substrates will slide
into the active site of the enzyme, and the enzyme will hold the substrates into position for bondforming. When an enzyme mediates molecule-breaking (catabolism), the substrate will fit into the
active site and be held in the appropriate position for bond-breaking (see below).
http://www.chemicalconnection.org.uk The enzyme’s three-dimensional shape is critical for its function – only the appropriate substrate
will correctly fit into the active site of an enzyme (see below).
http://www.chemicalconnection.org.uk Extremes of heat, pH and salt concentration can disrupt the three-dimensional shape of a protein.
When this happens, the protein is called “denatured.” The unfolded shape is usually permanent
(think of a fried egg white, or curdled milk).
Will a denatured enzyme be functional? ________________________________
Enzyme-mediated reactions are typically written with the substrate(s) or reactant(s) to the left of the
arrow, the enzyme name over the arrow, and the product(s) at the right of the arrow. For example,
the reaction in which the enzyme lactase digests the disaccharide lactose into its component
monosaccharides would be written as follows:
lactase lactose
glucose + galactose
1 (Modified from A. Goodwin)
Name: ______________________________
For an enzyme to work, it must encounter its substrate in the solution. One of the factors that can
affect how quickly an enzyme meets its substrate is temperature. With increasing temperature,
molecules move more quickly – and bump into each other more frequently. The benefit of
increasing temperature is only observed within the tolerance range of the enzyme, however –
when a temperature is high enough to disrupt the enzyme’s three-dimensional shape, function will
be lost.
Lactose Intolerance
Lactase is an enzyme that digests the milk sugar lactose into its monosaccharides, glucose and
galactose. Only monosaccharides can be transported from the small intestine into the
bloodstream. People who are lactose intolerant lose the ability to produce lactase – or produce
lower quantities of this enzyme – after the first few years of life. In general, the ability of an adult
to produce lactase is genetically determined – a mutation in the DNA region that controls lactase
production allows this protein to be produced into adulthood. Lactose-tolerant adults can stop or
reduce production of lactase following intestinal infection or for unknown reasons, however.
1. Which type of molecule is lactose? What is its structure?
2. Which type of molecule is lactase? What is its function?
3. Based on the presentation on lactose intolerance, why have certain ethnic groups evolved the
ability to digest lactose as adults?
4.
Look at the package of “Lactaid” tablets. What is the active ingredient?
5. Would drinking lactase-treated milk while eating ice cream be as effective as taking a Lactaid
tablet with the ice cream? Explain. (Hint: Remember that lactose digestion takes place in the
small intestine – after the stomach.)
2 (Modified from A. Goodwin)
Name: ______________________________
Catalase Activity
Catalase is an enzyme that breaks down hydrogen peroxide into water and oxygen.
catalase
H 2O 2
Hydrogen
peroxide
H 2O + O
water oxygen Since the product oxygen is actually released as pairs of oxygen, the reaction is better written as:
catalase
2H2O2
2H2O + O2
1. What is the substrate for catalase? ________________________
2. What are the products for this reaction? __________________________________
3. Why will bubbling be observed as the reaction proceeds?
4. Examine the hydrogen peroxide by itself in the
beaker. Why do you not see any bubbling?
http://biowithberkeley.blogspot.com/2007/10/apbiology-lab-2-enzyme-catalysis.html
5. Prepare a catalase solution by processing 100 ml water and ½ of a potato in a blender. Strain
through cheesecloth to remove the potato bits.
3 (Modified from A. Goodwin)
Name: ______________________________
Substrate Specificity
1. Label three test tubes with the numbers 1, 2 and 3.
2. Add 1 ml water to tube 1, and 1 ml catalase to tubes 2 and 3.
3. Add 4 ml hydrogen peroxide to tube 1. Swirl to mix and wait at least 20 seconds for bubbling
to develop. Measure the height of the bubble columns in millimeters (mm) and record your
results in Table 1 below.
4. Add 4 ml hydrogen peroxide to tube 2. Swirl, wait 20 seconds and measure the bubble
column height. Record your results in Table 1.
5. Add 4 ml sucrose solution to tube 3. Swirl, wait 20 seconds and measure the bubble column
height. Record your results in Table 1.
6. Explain why bubbling was observed – or not observed – for each tube and write your
explanation in Table 1.
Table 1. Substrate Specificity
Tube
Contents
1
water
hydrogen
peroxide
2
catalase
hydrogen
peroxide
3
catalase
sucrose solution
Bubble
column height
Explanation
7. Draw a diagram illustrating why bubbles are observed for one substrate, but not another.
4 (Modified from A. Goodwin)
Name: ______________________________
Effects of Enzyme Amount on Reactions
1. Label three test tubes with the numbers 1, 2 and 3.
2. Add 1 ml catalase to tube 1, 2 ml catalase to tube 2, and 3 ml catalase to tube 3.
3. Add 4 ml hydrogen peroxide to each tube; swirl, wait 20 seconds and measure the bubble
column height for each tube. Not that the bubble column may exceed the height of the tube;
estimate the total column height as best as possible. Record your results in Table 2 below.
4. Explain differing degrees of bubbling were observed for each tube and write your explanation
in Table 2.
Table 2. Enzyme Amounts
Tube
Enzyme volume
1
1 ml
2
2 ml
3
3 ml
Bubble
column height
Explanation
5. If unlimited time had been allowed, would the bubble column height have been the same for
all tubes? Explain.
6. If the substrate volume had been varied, would the bubble height have been the same for all
tubes given unlimited time?
5 (Modified from A. Goodwin)
Name: ______________________________
Effects of Temperature on Enzyme Activity
1. Label three test tubes with the numbers 1, 2 and 3, and an identifying mark for your group.
2. Add 1 ml catalase to each tube.
3. Place tube 1 in a cold water bath and tube 2 in a boiling water bath. Leave tube 2 at room
temperature. Wait for 10 minutes.
4. Add 4 ml hydrogen peroxide to each tube; swirl, wait 20 seconds and measure the bubble
column height for each tube. Record your results in Table 3 below.
5. Explain differing degrees of bubbling were observed for each tube and write your explanation
in Table 3.
Table 3: Temperature Effects
Tube
Water bath
1
Ice water
2
Room
temperature
3
Boiling
Temperature Bubble
(°C)
column height
Explanation
6. Graph the effects of temperature on bubble column height. Remember to put the variable that
you control – temperature – on the x (horizontal) axis and the variable that you measure –
bubble column height – on the y (vertical) axis.
6 (Modified from A. Goodwin)
Name: ______________________________
Effects of pH on Enzyme Activity
1. Label three test tubes with the numbers 1, 2 and 3.
2. Add 1 ml catalase to each tube.
3. Add water of varying pH to each tube: to tube 1 add 2 ml water with pH = 3; to tube 2 add 2
ml water with pH = 7; and to tube 3 add water with pH = 11. Note that the solutions with pH
2 and 11 are relatively strong acids and bases, respectively – if you spill these solutions on
your skin, rinse immediately with water.
4. Wait 1 minute.
5. Add 4 ml hydrogen peroxide to each tube; swirl, wait 20 seconds and measure the bubble
column height for each tube. Record your results in Table 4 below.
6. Explain differing degrees of bubbling were observed for each tube and write your explanation
in Table 4.
Table 4. pH and Enzymes
Tube
pH
1
3
2
7
3
11
Bubble column
height
Explanation
7. Graph the effects of temperature on bubble column height. Remember to put the variable that
you control – pH – on the x (horizontal) axis and the variable that you measure – bubble
column height – on the y (vertical) axis.
7 (Modified from A. Goodwin)
Name: ______________________________
Challenge:
You are employed by a company that would like to tap into the growing market of foods for
lactose-intolerant people. Since the optimal conditions for each enzyme are different, your job is
to identify the ideal environment for lactase activity. Explain how you would determine the
conditions that would allow you to digest the maximum amount of lactose in 1 hour. For each
condition, explain why it would be important to look at this factor, and draw a graph showing
hypothetical results.
Lactose/lactase concentrations
Temperature
pH
8