Download Properties of Enzymes Lab

Name: ____________________________________________ Period: ______ Date: _____________
Honors Biology
Factors that Effect Enzyme Function
Introduction: Enzymes are proteins that speed up chemical reactions by lowering the activation energy needed to start the
reaction. They do this by binding to the reactants (substrates) and changing shape which places the substrates in a position that
facilitates bond breaking or bond formation. The product(s) of the reaction form thousands of times more rapidly than they
would without the enzyme.
Every enzyme has a unique three-dimensional shape determined by its primary sequence of amino acids. The pocket in the
enzyme into which the substrate fits is called the active site. The active site of an enzyme complements the substrate in shape
and charge.
All enzymes share the following basic properties, each of which will we will study in the lab:
1. Enzymes are specific. Almost all enzymes will catalyze a single type of reaction. This is because the shape of the
enzyme must complement the substrate (reactants) perfectly in shape and charge.
2. Enzymes are not altered by the reactions they catalyze. They can be reused over and over again.
3. Enzyme activity is affected by the amount of available substrate. If a large concentration of substrate is present,
more enzymes can bind and catalyze the reaction creating more product.
4. Environmental conditions that alter the shape of the enzyme and its active site impair enzyme function. High
temperatures, acidic or salty conditions all disrupt the hydrogen bonds that maintain the tertiary structure of enzymes.
When this happens, the enzyme denatures (reverts to its primary structure). This alters or destroys the active site so
the enzyme can no longer bind the substrate and catalyze the reaction.
The enzyme you will be working with today is called catalase. Catalase can be found in the cells of all living things. It speeds
up the following reaction by bonding to H2O2 (the substrate).
H2O2
H2O
+
O2
catalase
Hydrogen Peroxide
Water
Oxygen
The amount of hydrogen peroxide broken down can be measured by the amount of oxygen released as a gas from the reaction.
In this lab, it will be measured by the amount of foam produced in the test tube.
Purpose: The purpose of this lab is to investigate each of the four properties of enzymes introduced in the background above.
Materials:
4 test tubes
stop watch
liver solution
Hydrogen Peroxide
Ruler
Graduated Cylinder
stirring rod
Directions:
1) Put on your goggles. Leave them on for the entire lab.
2) Take your test tube rack up to the front of the room. Place the following into each of your test tubes
Tube 1: 40 drops of fresh liver solution
Tube 2: 40 drops of fresh liver solution
Tube 3: 40 drops of fresh liver solution
Tube 4: 40 drops of acid treated liver solution
Part I: Enzyme Specificity
Test tubes one and two will demonstrate the property of enzyme specificity.
You will be adding the substrate H2O2 to test tube #1. Read all instructions before adding the H2O2.
3) Carry test tube #1 over to the sink. (This test will produce so much foam it will pour out of the test tube.) Use the H2O2
pipette to fill your graduated cylinder with 5 ml of hydrogen peroxide. Pour the H2O2 into tube #1. Measure the foam
production in mm with a ruler immediately and every 20 seconds until 60 seconds has passed. (If it overflows, measure
the distance from the top of the liquid to the tip of the test tube and make a > sign on the data table to show it was larger
than that number.) See figure 2 below.
4)
5)
6)
7)
8)
9)
Place test tube 1 back into the test tube rack. Do not dump it out. You will be using it again later in the lab.
Rinse your graduated cylinder out multiple times.
Use the water pipette to transfer 5 ml of water into the graduated cylinder.
Pour the water into the liver solution of test tube 2.
Use the ruler to measure the foam production immediately and every 20 seconds until 60 seconds has passed.
Record your data in the table.
Part II: Enzymes are not altered or used up by the reactions they catalyze.
Test tube #1 will be used again to demonstrate this property. Substrate was already added to the enzyme in this test
tube in a previous test. If the enzyme was not altered or used up in the reaction, we should be able to add more
substrate (hydrogen peroxide) and get the same reaction. Let’s try it.
10) Carry test tube #1 over to the sink. Use the H2O2 pipette to fill your graduated cylinder with 5 ml of hydrogen peroxide.
Pour the H2O2 into tube #1. Measure the foam production in mm with a ruler immediately and every 20 seconds until 60
seconds has passed. (If it overflows, measure the distance from the top of the liquid to the tip of the test tube and make a >
sign on the data table to show it was larger than that number.)
11) Record your data in the table.
Part III: Enzymes are regulated by the amount of available substrate.
Test tube #3 will be used to demonstrate this property. You are going to lower the concentration of available substrate
for the liver in test tube #3.
12) Use the H2O2 pipette to put 5 drops of hydrogen peroxide into the graduated cylinder. Use the water pipette to add
enough water for a total volume of 5 ml. Pour the diluted H2O2 into test tube #3. Measure the foam production in mm
with a ruler immediately, and every 20 seconds until 60 seconds has passed. Record in the data table.
Part IV: Factors that alter the shape of the enzyme and its active site impair enzyme function.
Test tube #4 has liver solution that was exposed to HCl (pH of 1). You will add standard strength H2O2 to this tube to
determine what effect the acid has on the enzyme’s function.
13) Use the H2O2 pipette to fill your graduated cylinder with 5 ml of hydrogen peroxide. Pour the H2O2 into test tube #4.
Measure the foam production in mm with a ruler immediately, and every 20 seconds until 60 seconds has passed. Record
in the data table.
14) Clean-up. Rinse all test tube solutions down the sink with running water. Wash all tubes, the test tube rack, the ruler, and
the graduated cylinder with soapy water. (Anything that came into contact with liver must be washed.) Wash off your lab
station. Dry everything with a paper towel. Reline your plastic tray with a fresh paper towel. Call your instructor over for
an inspection.
Complete the analysis and conclusion of the lab.
Name: ______________________________________________________________________ Period; _________
Honors Biology
Properties of Enzymes Lab
Pre-Lab Questions:
1. What is the name of the enzyme in this lab? Where are we obtaining the enzyme from?
2.
What substrate does the enzyme work on?
3.
Write the chemical reaction the enzyme catalyzes. Place the name of the enzyme over the arrow in the chemical
reaction. Use brackets to identify the reactants and products.
Data: Foam Production Generated in Test Tubes with Liver and Hydrogen Peroxide Solutions
Foam Production (mm)
Tube #1 (fresh
Tube #2 (fresh
Tube #1 (fresh
Tube #3 (fresh
liver + standard
liver + water)
liver + standard
liver + reduced
Time (seconds)
concentration
concentration
concentration of
H2O2 trial #1)
H2O2 trial #2)
H2O2)
Tube #4 (acid
treated liver +
standard
concentration
H2O2)
1 second
20 seconds
40 seconds
60 seconds
Analysis: Read the Background of the lab to answer these questions.
1. What was the purpose of adding water to the liver solution in test tube #2 compared to the hydrogen peroxide that
was added in test tube #1? (What property of enzymes were we trying to demonstrate?)
2.
What relationship exists between the shape and charge of an enzyme active site and its substrate?
3.
Was foam produced the second time hydrogen peroxide was added to the liver in test tube #1? What property of
enzymes does this demonstrate?
4.
What effect did lowering the concentration of the hydrogen peroxide have on the enzyme function? Why?
5.
What did the acid do to the shape of the enzyme? (Provide the term and description of what this means in your
answer.)
6.
What effect did the acid have on the function of the enzyme? Why?