Enzymes Adapted from Air All Around: Oxygen Investigation

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1 Enzymes Adapted from Air All Around: Oxygen Investigation Author: Doris Pun & Brittland DeKorver Institute for Chemical Education and Nanoscale Science and Engineering Center University of Wisconsin-Madison Purpose: To learn about enzymes Learning Objectives: 1. Learn what an enzyme is and what it does. 2. Observe an enzyme in action. 3. Learn how enzymatic activities can be altered. Next Generation Science Standards (est. 2013): PS1.A: Structure and Properties of Matter (partial) PS1.B: Chemical Reactions PS3.B: Conservation of Energy and Energy Transfer (partial) National Science Education Standards (valid ): Standard B: Physical Science o Properties and changes in properties of matter o Transfer of energy Standard C: Life Science o Structure and function of living systems o Regulation and heredity Standard D: Science and Technology o Abilities of technological design o Understanding about science and technology Standard F: Science in Personal and Social Perspectives o Science and technology in local, national, and global challenges Suggested Previous Activity: Green Chemistry: Catalysts Grade Level: 5-8 Time: 60 minutes Materials: coins/caps Timer Tennis ball or similar sized ball Safety glasses Gloves 3 % hydrogen peroxide (H2O2) 13 ml test tubes

2 Cut potatoes (0.5 x 0.5 x 2 cm sticks) o Raw, boiled, frozen Dilute acid (0.1M HCl) Dilute base (0.1M NaOH) Pipettes CuSO4 Spatula Apples Knife Petri dishes Lemon juice Microscopes Microscope slides of different enzymes Yeast Safety: Wear gloves and safety goggles while handling the peroxide, liquid nitrogen, acid and base solutions. Preparation ahead of time: Prepare small bottles of 3 % H2O2 for student use. Cut the potatoes into approximately identical sizes, 0.5cm x 0.5cm thick and 2-3cm long. Boil 1/6 of the potatoes. Definitions: - Enzyme: catalyst that helps perform a chemical reaction within living cells/organisms - Substrate: the molecule that the enzyme alters - Product: the molecule that the enzymes produce from the substrate - Active site: site within the enzyme that the substrate binds to and the chemical reaction occurs - Inhibitor: molecules that bind to the active site of an enzyme in such as way that prevents substrates from binding Introduction: Catalysts are substances that speed up a chemical reaction without being consumed in the reaction, making it reusable. Catalysts are used to make many plastics and medicines, and biological catalysts (enzymes) perform all sorts of important reactions in our bodies. They are usually made up of large proteins or polymer chains. In a typical enzyme-catalyzed reaction, a specific molecule called the substrate binds to the enzyme in a pocket specifically designed for the molecule called the active site. The enzyme then converts the substrate into one or multiple products and releases the product. The enzyme can then bind another molecule and repeat the reaction. Any changes to an enzyme s active site will affect the catalytic efficiency of the enzyme. A common change is the denaturing of the proteins, where the shape and structure of the proteins are lost. A familiar example is the denaturing of egg whites when cooking/heating. Another familiar example is the addition of vinegar (an acid) to milk causing it to curdle. These examples demonstrate that the temperature and acidity can cause denaturation. The enzymatic efficiency can also be affected by the presence of inhibitors. Inhibitors bind to the active site,

3 stopping the substrate from entering the active site so that the chemical reaction can occur. Many drugs are actually inhibitors, working by inhibiting enzymes in bacteria, viruses, or cancerous cells. Tell the students that today they are going to learn about enzymes and they are going to explore several enzymes in action. They will also learn how to alter an enzyme s activity. Procedures: 1. Enzyme Models fingers catalyzed reaction (Group activity) a. Special role - rotate amongst the students i. Timer will measure how long the reaction takes to reach completion b. Explain to the students that the reaction is flipping all the caps/coins such that they are all either heads/bottoms up c. Normal enzymatic reaction: (perform 4-10 times total) i. Explain that the students are enzymes and can use all their fingers to do the reaction ii. Time and record on a table datasheet how long it takes for student #1 to flip all heads up iii. Repeat step iii with student #2 for tails up iv. Repeat steps iii-iv at least another round with different students v. Calculate the average enzymatic reaction completion time d. Partial denaturing/decomposition of enzyme reaction i. Explain that the enzymes are now slowly decomposing either by high temperatures, acid or base contact, etc. ii. Now the students are only allowed to use their 2 index fingers to do the reaction iii. Repeat Steps c. ii-v. e. Enzyme inhibition reaction select only one student to perform i. Explain that the enzymes will now be introduced to inhibitors ii. Place a tennis ball into the palms of each enzyme s hand iii. Repeat Steps c. ii & v. f. Discuss and compare the different conditions of each reaction along with their reaction rates. 2. Lemon Juice Inhibition of Apple Browning a. Have an adult slice up enough slices to give each student 2 small slices b. Put slices on separate Petri dishes and clearly label one as lemon and the other as no lemon c. Squeeze and rub lemon juice on the lemon slice d. Set aside for 15 minutes and observe any differences between the two slices

4 3. Potato (enzyme = catalase) Catalyzed Decomposition of Peroxide (H2O2) into Water and Oxygen wear safety goggles and gloves This activity should be familiar to the students who performed the manganese dioxide, MnO2, catalyzed peroxide decomposition reaction in Green Chemistry: Catalysts. a. Fill a test tube half way with peroxide, H2O2 b. Carefully drop a small stick of raw potato into the test tube c. Record any observations, ie. bubbling, temperature of reaction? d. Repeat with different conditions to try to inhibit/denature enzyme. Suggestions: i. Boiled potato stick ii. Frozen potato stick iii. With dilute acid (0.1M HCl) iv. With dilute base (0.1M NaOH) 1. tip: add acid/base to potato first v. With copper sulfate, CuSO4, as inhibitor 1. tip: measure out mg CuSO4 2. coat potato with as much of the measured out CuSO4 3. put rest of the CuSO4 in the test tube of peroxide and mix 4. drop CuSO4-coated potato in the test tube 4. (Optional if microscope available) Microscope Activity a. Look at different prepared enzymes on slides b. Observe yeast or potatoes with drop of peroxide Discussion: Ask students if they are familiar with hydrogen peroxide. Tell them that it is commonly found in medicine cabinets. It works as an antibacterial agent as it can destroy cells. It is also a common byproduct in metabolism in living organisms. Tell them that hydrogen peroxide has two oxygen atoms in each hydrogen peroxide molecule, and that the molecules break down. The atoms rearrange into oxygen gas and water. The process will happen faster if an enzyme, like catalase, is used. Catalase is an enzyme found in most organisms to rapidly perform this decomposition reaction by binding peroxide and releasing oxygen and water. High concentrations of catalase are found in the liver. Potatoes are the source of catalase in this lesson. The boiling, lowered and elevated acidity by adding acid and base, respectively denature the enzyme and lower the enzyme s efficiency. The freezing of the enzyme only denatures the enzyme partially. The copper sulfate serves as a noncompetitive inhibitor to catalase, binding to the enzyme s active site and not allowing for peroxide to bind. Apples and bananas contain an enzyme called catechol oxidase which catalyzes the reaction of the compound, catechol, and oxygen to a new molecule called benzoquinone, which is orange in color and also toxic to bacteria, slowing down the spoilage of the fruit. Adding lemon juice, which is an acid, lowers the acidity

5 of the apple and denatures the catechol oxidase enzyme, preventing the transformation, which causes the orange-brown color on apples. Evaluation: What is a catalyst? Are catalysts usually consumed in a chemical reaction? What is the relationship between catalysts and enzymes? Do you think your body has some enzymes? Why or why not. This lesson is the product of the Institute for Chemical Education and the Nanoscale Science and Engineering Center at the University of Wisconsin-Madison. This Material is based upon work supported by the National Science Foundation under grant number DMR SCIENCountErs Lessons are licensed under a Creative Commons Attribution- NonCommercial 4.0 International License. Permissions beyond the scope of this license may be available by ing ice@chem.wisc.edu.