Description *Lab is not needed to complete this assignment* You are simply making notes based on fa ...
Description *Lab is not needed to complete this assignment* You are simply making notes based on facts as in what will happen in the situations. Answer all questions and make notes for all statements. Background Figure 1. A Pineapple Pineapples contain a combination of protein-digesting enzymes called bromelain. Why Does It Sometimes Sting When You Eat Pineapple? Pineapple (Figure 1) contains bromelain, a mixture of two protein-digesting enzymes. When you eat pineapple, the enzymes start to digest the proteins in your tongue and mouth, causing a stinging sensation. However, there are not enough enzymes in pineapple to damage your mouth, just enough to make it sting. undefined Bromelain is used as a meat tenderizer because the enzymes break down the proteins in the meat, making it softer. However, you should be careful when mixing bromelain or pineapples with meat. If you leave the mixture too long, the enzymes will break down many of the proteins in the meat and it will become mushy and unappealing. Enzymes Enzymes are biological catalysts, or molecules that promote biochemical reactions without being changed or used up in the process. Enzymes can start or speed up a reaction. For example, in the presence of certain enzymes, a reaction can proceed at the rate of millions of times per second! In the absence of the enzymes, however, the reaction could take years to complete, if it finished at all. When an enzyme increases the rate of a reaction without being altered itself in the end, it is called catalysis. undefined Most enzymes are globular proteins with a specific 3-D shape that is determined by the electrostatic charges of the enzymes’ molecules attracting and repelling one another. The 3-D shape of a protein is critical to its function because it creates the enzyme’s active site. The active site is a pocket in the enzyme where the substrate, or molecule undergoing catalysis, binds. undefined Once a substrate encounters the active site of an enzyme, the enzyme is induced to change shape slightly to better accommodate the substrate (Figure 2). This is known as the induced fit model. The enzyme then facilitates a chemical reaction that changes the substrate into products. When the process is complete, the enzyme releases the products and is ready to begin the process all over again with a new substrate molecule. Thus, enzymes are reusable and not exhausted while facilitating the reaction. Figure 2. Induced Fit Model of Enzyme Activity Environmental Factors Affecting an Enzyme’s Ability to Catalyze Reactions PH An enzyme’s 3-D shape, and therefore functionality, is affected by pH. In the presence of excess H+ ions, which cause an acidic condition, or OH- ions, which cause an alkaline condition, the active site on the enzyme becomes progressively distorted as ionic interactions between the protein molecules shift. This decreases the enzyme's activity until it can no longer function as a catalyst. This process is called denaturation of the enzyme. Most enzymes work in a range of pH values, but function best at their optimum pH. TEMPERATURE Temperature also affects the activity of enzymes. Catalysis occurs at a faster rate as temperature increases, until it reaches its optimum temperature. Each enzyme has an optimum temperature, beyond which: the enzyme's hydrogen bonds and hydrophobic interactions weaken the functional 3-D shape of the enzyme is lost catalysis decreases Boiling temperatures will denature most enzymes by stretching the molecular bonds present in the enzyme beyond repair. Most enzymes work in a range of temperatures, but function best at their optimum temperature. SUBSTRATE CONCENTRATION An increase in substrate concentration will increase the rate of the chemical reaction until there is enough substrate to saturate all of the enzyme molecules. When this happens, the reaction rate will level off because the rate can only go as fast as the number of enzymes facilitating the reactions (Figure 3). In a closed reaction where substrate supply is finite, the reaction rate will eventually decrease as the substrate supply starts to run out until there is no substrate left, and thus no new products created. Figure 3. Example of Substrate Concentration Effect on Reaction Rate: Assuming there is an unlimited supply of substrate, the reaction rate will level off as soon as all the enzyme molecules are saturated with substrate. ENZYME CONCENTRATION As the enzyme concentration increases, the reaction rate will also increase, as long as there is a supply of substrate molecules (Figure 4). This is possible because enzyme molecules are not used up during a reaction like substrate molecules are. The enzymes can immediately begin catalyzing a new reaction as soon as they finish a reaction. Figure 4. Example of Enzyme Concentration Effect on Reaction Rate: Assuming there is an unlimited supply of substrate, the reaction rate will continue to increase as the enzyme concentration increases. As soon as an enzyme molecule finishes catalyzing one substrate molecule to its products, it can perform the reaction again on another substrate molecule. Enzymes are not limiting of reaction rate as long as they have substrate to work on. About This Lab In this lab, you will examine the effects of temperature, pH, and substrate concentration on the enzyme, catalase. Catalase greatly accelerates the decomposition of dangerously reactive hydrogen peroxide into harmless water, oxygen, and heat as shown in the equation below. undefined 2H2O2??????catalase2H2O+O2+heat undefined Because oxygen gas is created as a product, the rate of the reaction is easy to evaluate by looking at how much of and how quickly the gas is produced. undefined Most living cells contain catalase because many cellular reactions form hydrogen peroxide as a product. Mammalian liver has a high concentration of catalase and is the source of the catalase used in this lab. EXPERIMENTS Open the simulation by clicking on the virtual lab icon below. The simulation will launch in a new window. You may need to move or resize the window in order to view both the Procedure and the simulation at the same time. Follow the instructions in the Procedure to complete each part of the simulation. When instructed to record your observations, record data, or complete calculations, record them for your own records in order to use them later to complete the post-lab assignment. Procedures Experiment 1: Effect of Temperature on Catalase Activity Take a constant temperature bath, thermometer, and gas syringe from the Instruments shelf and place them onto the workbench. Set the temperature bath to 10 °C. Take a small test tube from the Containers shelf and place it in the constant temperature bath. Add 5 mL of 3% hydrogen peroxide from the Materials shelf to the test tube. Add 5 mL of water from the Materials shelf to the test tube. Move the thermometer from the workbench into the test tube. Wait until the temperature stabilizes at the same temperature as the bath. Move the gas syringe from the workbench into the test tube. Click the gas syringe and note the initial volume of gas in the syringe. Add 1 mL of catalase solution from the Materials shelf to the test tube and immediately note the time on the lab clock, particularly the seconds. After 15 seconds elapse, click the gas syringe again to determine the volume of gas released from the reaction. Record the two volumes (initial and after 15 seconds) to reference later. Move the thermometer and gas syringe to the workbench. Empty the test tube in the waste bin, and place it in the sink. Repeat steps 2 – 13 four additional times, using temperatures of 21.5, 40, 60, and 80 °C instead of 10 °C. Record the optimum temperature for catalase activity based on your results. Clear the workbench by dragging instruments back to the Instruments shelf and by emptying containers in the waste bin and then placing the empty containers in the sink. Experiment 2: Effect of Substrate Concentration on Catalase Activity Take four constant temperature baths off the Instruments shelf and place them side to side onto the workbench. Space is limited, so place them as close to the left wall and each other as possible. Set all four constant temperature baths to the optimum temperature for catalase activity that you determined in Experiment 1. Take four small test tubes from the Containers shelf and place one into each constant temperature bath. Double-click on each test tube and rename them: 1, 2, 3, and 4. Take four thermometers from the Instruments shelf and place one into each test tube. Using items found on the Materials shelf, fill the four test tubes as follows. undefined Test Tube Water (mL) 3% Hydrogen Peroxide (mL) 1 8 2 2 5 5 3 2 8 4 0 10 undefined Wait until the temperature stabilizes and the tubes reach the same temperature as the bath. Take four gas syringes from the Instruments shelf and place one in each test tube. Click the gas syringe in test tube 1 and note the initial volume of gas in the syringe. Add 1 mL of catalase solution to test tube 1 and immediately note the time on the lab clock, particularly the seconds. After 15 seconds elapse, click the gas syringe in test tube 1 again to determine the volume of gas released from the reaction. Record the two volumes (initial and after 15 seconds) to reference later. Repeat steps 8 – 10 for the three other test tubes. Record the concentration of hydrogen peroxide that produced the most gas. Clear the workbench by dragging instruments back to the Instruments shelf and by emptying containers in the waste bin and then placing the empty containers in the sink. Experiment 3: Effect of pH on Catalase Activity Take three constant temperature baths from the Instruments shelf and place them onto the workbench. Set all three constant temperature baths to the optimum temperature for catalase activity that you determined in Experiment 1. Take three small test tubes from the Containers shelf and place one into each constant temperature bath. Take three thermometers from the Instruments shelf and place one into each test tube. Add 5 mL of 3% hydrogen peroxide from the Materials shelf to each test tube. Add 5 mL of pH 2 buffer from the Materials shelf to the first test tube. Double-click on this test tube to rename it “pH 2 buffer”. Add 5 mL of pH 6 buffer from the Materials shelf to the second test tube. Double-click on this test tube to rename it “pH 6 buffer”. Add 5 mL of pH 10 buffer from the Materials shelf to the third test tube. Double-click on this test tube to rename it “pH 10 buffer”. Wait until the temperature stabilizes and the tubes reach the same temperature as the baths. Take three gas syringes from the Instruments shelf and place one in each test tube. Click the gas syringe in the pH 2 buffer test tube and note the initial volume of gas in the syringe. Add 1 mL of catalase solution from the Materials shelf to the pH 2 buffer test tube and immediately note the time on the lab clock, particularly the seconds. After 15 seconds elapse, click the gas syringe in the pH 2 buffer test tube again to determine the volume of gas released from the reaction. Record the two volumes (initial and after 15 seconds) to reference later. Repeat steps 11 – 13 with the pH 6 buffer and pH 10 buffer test tubes. Record the optimum pH for catalase activity. Clear the bench of all materials, containers, and instruments, then return to your course page to complete any assignment for this lab.
