Experiment 2: Effect of Temperature on Enzyme Activity *Photo of Experiment Results Required Data Table 2: Balloon Circumference vs. Temperature Tube | Temperature (°C) | Balloon Circumference (Uninflated; cm) | Balloon Circumference (Final; cm) 1 – (Cold) | 10 °C | 11.5 cm | 13.5 2 – (RT) | 22 °C | 11.5 cm | 23.0 cm 3 – (Hot) | 38 °C | 11.5 cm | 14.2 Post-Lab Questions: 1. What reaction is being catalyzed in this experiment? Write the equation for the reaction below: -----------> + 2. What is the enzyme in this experiment? What is the substrate? Catalase is the enzyme. The substrate is hydrogen peroxide 3. What product was measured in this experiment to determine enzyme activity and the rate of the reaction? 4. How did the colder temperature affect enzyme function? Why do you think the cold temperature had this effect? Hint: Molecules move at a slower speed at low temperatures. The cold temperature produced a slower enzyme activity rate because the molecules move slowly due to the reduction of frequency of enzyme-substrate collisions. 5. Predict what would happen to enzyme activity and reaction rate if you changed the following conditions in this experiment. a. Increased the temperature over 100 °C: The enzyme activity would increase breaking the bonds in hydrogen peroxide or denaturing the enzymes in yeast. b. Increased the amount of substrate: c. Lowered the pH: d. Increased salt concentration: 6. Could the enzyme catalase speed up another type of reaction such as digestion of lactose? Why or why not? Match the following words with their definitions. A. Product B. Active site C. Enzyme D. Substrate E. Activation energy 7. Amount of energy required to initiate a chemical reaction; lowered by a catalyst. 8. Protein catalyst that speeds up a chemical reaction without being consumed. 9. Reactant that enzyme act upon. 10. Region on the surface of an enzyme where substrates bind. 11. Substance formed from substrate(s) at the end of a chemical reaction.
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Experiment 2: Effect of Temperature on Enzyme Activity Table: Balloon Circumference vs. Temperature Post-Lab Questions: 1. What reaction is being catalyzed in this experiment? Write the equation for the reaction below: 2. What is the enzyme in this experiment? What is the substrate? 3. What product was measured in this experiment to determine enzyme activity and the rate of the reaction? 4. How did the colder temperature affect enzyme function? Why do you think the cold temperature had this effect? Hint: Molecules move at a slower speed at low temperatures. 5. Predict what would happen to enzyme activity and reaction rate if you changed the following conditions in this experiment: a. Increased the temperature over 100 °C: b. Increased the amount of substrate: c. Lowered the pH: d. Increased salt concentration: 6. Could the enzyme catalase speed up another type of reaction such as digestion of lactose? Why or why not? Match the following words with their definitions: A. Product B. Active site C. Enzyme D. Substrate E. Activation energy 7. Amount of energy required to initiate a chemical reaction; lowered by a catalyst. 8. Protein catalyst that speeds up a chemical reaction without being consumed. 9. Reactant that enzyme act upon. 10. Region on the surface of an enzyme where substrates bind. 11. Substance formed from substrate(s) at the end of a chemical reaction.
Madhur L.
Isolation and Characterization of Enzymes Objectives After completing this exercise students should be able to do the following: 1. Understand how enzymes work. 2. Explain the effect of varying enzyme concentration on the activity of enzymatic reaction. 3. Explain the effect of varying temperature and pH on enzymatic activity. 4. Understand the effect of inhibitors on enzymatic activity. 5. Distinguish between competitive and noncompetitive inhibitors. INTRODUCTION Enzymes are biological catalysts that speed up the rate of chemical reactions in cells. The reactant that binds to the active site of enzyme is called a substrate. They together form a transient enzyme-substrate complex (E-S complex) which reduces the activation energy requirement of a reaction. Enzymes are effective in small amounts because they are not used up in the reactions that they catalyze. Since most of the enzymes are made of proteins, factors that affect protein structure such as temperature, pH, and various salts can also affect the activity of enzymes. The temperature and pH at which that an enzyme works best are called its optimal temperature and optimal pH respectively. The enzyme to be studied in this exercise is peroxidase. This enzyme is commonly found in a variety of cells. Peroxidase catalyzes the breakdown of hydrogen peroxide with the production of water and oxygen. If a reduced compound is present, it may be oxidized during this reaction by the enzyme. Certain dyes will go from a colorless to a colored form upon oxidation, a property we will take advantage of in detecting peroxidase activity. In the presence of hydrogen peroxide and peroxidase, orthophenylenediamine (OPD) will be changed from a colorless dye to a yellow compound as it is oxidized. The overall reaction may be described as follows: H2O2 + OPD -> H2O + OPD (colorless) (yellow) (reduced form) (oxidized form) Using a spectrophotometer, we can accurately quantitate the enzyme reaction by measuring...
While working with an enzyme in the lab to complete several different experiments you notice that the rate of your reaction speeds up as the temperature increase. So, in order to speed up the rate of your reactions for your next patch of experiments, you keep increasing the temperature of your enzyme solution. Eventually, you notice that your reactions are taking way longer to complete. Why did your reaction rate speed up and then slow down? (pick the best answer). As the temperature of the reaction increase the speed at which the enzymes moved increased. However, once the enzymes got too hot they began to denature and break down. In the first set of experiments there where too many enzymes for the number of substrates. Then as the enzymes got used up there was just the right amount of enzymes to substrates forming a perfect ration. Then at the end the reaction slowed down because all of the enzymes got used up. Temperature is one of the key factors that affect the rate at which reactions occur. As you increased the temperature the reaction should only speed up. If the reaction is slowing down that must mean that it is cooling down. As the reaction ran the enzymes bound with the substrates producing products. These products must have acted as inhibitors in some way to slow down the enzyme substrate reaction.
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