Protease Lab

Temperature is a measure of kinetic energy. As this movement increases, collision rate and intensity, and therefore reaction rates, increase. This experiment was conducted to determine if there is a minimum temperature that increase kinetic energy and denature enzymes to slow enzymatic reactions or fail to catalyze them. The experimental results indicate an increase in temperature will increase reaction rates until proteins denature.

• Fourthly, we kept the temperature at a constant 25°C using a water bath. At low temperatures, an increase in temperature causes an exponential increase in enzyme activity. This is because an increase in temperature provides more kinetic energy for the collisions of enzymes and substrates, so

The temperature can have a major impact on an enzyme. According to Campbell Biology author Reece etc. 2011 “The enzyme reaction will increase as the temperature increase with the increasing temperature….substrates collide with active sites more frequently when the molecules move rapidly.”(Reece etc 2011) Every enzyme hits its optimal temperature the reaction will be at its highest point.(Reece etc. 2011) When the

-VariablesoIndependent:The temperature of the milkoDependent:The time taken for the milk to solidifyoControlled:The same amount and type of milk usedThe same amount and concentration of enzyme mixture usedThe same test tube sizeResults:-TableAmount of enzyme mixture (mL)Amount of milk (mL)Temperature (oC)Time for milk to clot (min)Ex: 1 Ex: 2 Average2.551060+60+60+2.552034.2036.0035.12.55303.554.203.882.55402.102.252.182.55505.004.454.73Discussion:The experiment showed that changing the temperature did affect the rate at which the milk solidified. At low temperatures of 10oC and 20oC the milk took the longest to solidify and at 10oC did not even go lumpy after an hour. As the temperature increased the speed at which it reacted got faster until it reached around 40oC where the speed began to drop.

As the temperature increases, so will the rate of enzyme reaction. However, as the temperature exceeds the optimum the rate of reaction will decrease.

Question: How does changing enzyme concentration or temperature affect the reaction time of enzyme activity?

These results show how temperature of extreme high, or low affects enzyme activity. The highest rate of enzyme activity occurred at 37 Cº. Anything that was hotter or cold than 37 Cº slowed the reaction rate. As I thought, 100 degrees would denature the enzyme, and that was the case. The data provided shows exactly what temperatures enzymes work best, and worst. The objective was achieved as we discovered the different reaction rates under different temperatures. The results are reliable, as we know enzymes do not work well when under extreme heat or denaturation occurs. What I learned in this experiment was that enzymes don’t work well under cold temperatures because they tend to move slower. My hypothesis did not quite match, because I thought they work best at lower temperatures.

The hypothesis tested in this experiment was, if the temperature of enzyme catalysis were increased, then the reaction rate would increase, because enzyme-catalysis reacts by randomly colliding with substrate molecules, and the increase in temperature increases the speed of collision or reaction rate. The final data collected for the experiment was positive with my hypothesis. The coffee filter, covered in potato solution, sank and rose at a faster pace in the hydrogen peroxide when the temperatures were raised.

Also, large substances such as, proteins are broken down by enzymes called proteases into amino acids, which is used and stored as energy. An example of this is; when the body gets exhausted the fuel which is converted from the protein is used as energy and also to repair worn out tissues to keep the body healthy and energised. It also stimulates the immune system and aids the body in ATP production.

Between 36.7°C and 4° C there was a decrease in reaction rate. This is majorly due to the fact that less enzyme-substrate complexes are formed as the kinetic energy is decreased and both the catalase and hydrogen peroxide have less collisions with one another (“The Kinetic Molecular Theory”). A temperature decrease would have also caused the molecules to be more tightly packed together, thereby reducing the flexibility of catalase, making enzyme-substrate complexes less likely to occur.

However if the temperature exceeds the optimum temperature the enzyme becomes denatured. This is because there is too much energy causing the enzyme molecules to vibrate causing the bonds maintaining their tertiary structure to break. The enzyme unravels causing the shape of the active site to change so it can no longer fit with the substrate.

Temperature: Temperature is an extremely important variable to consider, as it can affect the rate of reaction in a number of ways. If the temperature of a system is increased, more molecules will reach the activation energy and the rate of reaction will increase. The number of collisions will also increase. As enzymes must collide with substrates, an increase in temperature and thus kinetic energy will result in more collisions occurring in a given time. Increasing the temperature will also increase the heat of the molecules.

As illustrated in Table 1, increase in temperature resulted in the increase of the rate of catalase activity to a certain range. After the range, the temperature would continue to increase, but the rate of catalase activity will decrease significantly. Rate of enzyme activity increased as temperature increased is supported by the Collision Theory. The collision theory states in order for a reaction to occur, reacting particles must collide with one another (McGraw-Hill Ryerson Chemistry 12). Particles must collide with sufficient energy to overcome activation energy and with the correct geometrical orientation. Increase in temperature and rate of catalase activity, increases kinetic energy of hydrogen peroxide molecules. This allows a faster movement of hydrogen peroxide molecules increasing the collision frequency and the effectiveness of the collisions. As a result, more reactions will be speed up and increase catalase activity. The range where the rate of catalase activity is the highest is at 60°C. This would be the peak for the catalase. At 60°C, the most collisions, more effective collisions take place and hydrogen bonds are loose making it easier for catalase to act on hydrogen peroxide. As a result of this 60°C is the temperature peak for the catalase. After 60°C, the rate of catalase activity increases. This is because enzymes are very fragile macromolecules. After a certain temperature range, intramolecular bonds are broken due to amino acids vibrations. This leads to the enzyme denaturing. Denaturing is when the structures in the enzyme has been altered and cannot be

reaction rate increases. If the temperature of an enzyme gets to high the reaction rate will slow

Treatment of cells with Pi1 a proteasome inhibitor, protects cells from ricin except when incubations are long. Similar effects are seen with an alternative proteasome inhibitor ALLN (Figure 1A/B). In contrast inhibiting endosomal proteases made little or no difference (Figure 1B).