Investigating Enzyme Activity
Investigating Enzyme Activity
Enzymes exist in all living things. They are composed of polymers of amino acids and are produced in living cells. Each cell contains several hundred enzymes, which catalyse a vast number of chemical reactions. Enzymes are known as 'biological catalysts' as they dramatically increase the rate at which reactions occur within living organisms, without being 'used up' or effecting the reaction in any other way. Enzymes catalysis saves the need for an increase in temperature in order to speed up reactions within living things. Such an increase in temperature would be lethal to the organism.
In this investigation I intend to explore the one of the factors that affect the rate of enzyme catalysis. My research from textbooks and the Internet suggests that this depends on several factors; temperature, pressure, pH and concentration. After research and careful consideration, I have decided to first look at how a change in temperature could affect the rate of reaction.
In order to design a suitable experiment and make a credible prediction, I must first explore more closely how temperature is likely to affect the rate of catalysis. Enzymes are specific - they only control one type of reaction; therefore I must use one specific enzyme in my experiment, in order to find a clear way of measuring the rate of reaction. Although they are specific, all enzymes work in a very similar way and have similar properties. They are all globular proteins and are all biological catalysts, they increase the rate of a given reaction without being used up and their presence does not change the nature of the reaction or the end product. Enzymes work by having an active site, made from amino acids. Here, substrate molecules will bind with the enzyme (and other substrate molecules if necessary) and a reaction takes place. The enzyme itself is not affected and releases the new chemical after the reaction. After release of the end product, more substrate molecules can bind with the active site.
Enzymes can catalyse anabolic reactions or catabolic reactions (involved in breakdown). In a catabolic reaction, the reverse would happen.
Using the information gained here together with my knowledge of kinetic theory, it is possible to understand how temperature affects the rate of reaction. Kinetic theory states that when a substance is heated, energy is given to the particles and they speed up. Therefore when heat is applied to an enzyme and substrate, the particles speed up, increasing the rate at which they bind with each other. This would suggest that the rate of reaction should increase as ...
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Enzymes can catalyse anabolic reactions or catabolic reactions (involved in breakdown). In a catabolic reaction, the reverse would happen.
Using the information gained here together with my knowledge of kinetic theory, it is possible to understand how temperature affects the rate of reaction. Kinetic theory states that when a substance is heated, energy is given to the particles and they speed up. Therefore when heat is applied to an enzyme and substrate, the particles speed up, increasing the rate at which they bind with each other. This would suggest that the rate of reaction should increase as the temperature is increased. This is not quite true, as there is a limit to the temperature at which an enzyme can work because excessive heat causes an enzyme to become denatured and stop working. Also, there is a minimum temperature at which an enzyme can function. Every chemical reaction requires activation energy in order to get started. Although enzyme catalysis greatly reduces this, some energy is still required. Because of this the reaction is still unable to happen below a given temperature (this varies depending on the type of enzyme and reaction, as does the maximum temperature). If warmed to above the activation temperature, an enzyme will work again as normal. A denatured enzyme, however, is damaged and will not work again even if cooled below the optimum temperature.
Prediction
I predict that the rate of reaction will increase as the temperature increases until the reaction reaches an optimum temperature. Above this optimum temperature, the rate of reaction will fall to zero very quickly, as the enzyme denatures. I must now conduct an experiment to test my prediction. I will do this using the enzyme 'catalyse'. Catalyse is found in most living organisms. It speeds up the catabolic reaction, which breaks down hydrogen peroxide into oxygen and water.
Apparatus
Using my information on catalyse, it is clear that one of the products of the reaction is oxygen. Therefore to measure the rate of reaction, I could measure the rate at which oxygen is produced. For this experiment I will need:
* Celery as the source of catalyse
* Hydrogen peroxide
* A water bath in which I can heat both the enzymes and substrate
* Thermometers to ensure both liquids are at the correct temperature
* Measuring cylinders in order to measure the amount of hydrogen peroxide used
* A stopwatch to enable me to work out the rate at which oxygen is produced
* A basin of water
* Conical flask in which the reaction will take place
* Bung and delivery tube
Method
To test my prediction I will heat the catalyse and hydrogen peroxide to a given temperature and allow them to react in the conical flask, by stirring the solution three times, starting the timer at the beginning of the reaction. The oxygen given off will pass through the delivery tube and bubble up into the test tube, which will be set full of water in a basin. I will allow the reaction to continue until the test tube full of water has been replaced by oxygen. The results will then be recorded in a table after the experiment has been conducted at a satisfactory amount of temperatures. My preliminary experiment suggests that suitable quantities to use would be 30cm³ hydrogen peroxide and 5cm³ of celery. I am now able to perform the experiment at a range of temperatures between 0° and 60°. My research suggests that most enzymes become denatured before 60°.
Fair Testing
To make sure this experiment is a fair test, the only factor I must vary must be temperature. This means that I have to keep the concentrations of the substance constant throughout the experiment. In order to ensure accuracy, I will conduct the experiment three times for each temperature and take a mean result to plot on the graph. The exact quantities used must first be determined by a preliminary experiment.
Results
Temperature (°C)
Time Taken (average)
0°
2 minutes 40 seconds 26 ms
20°
minute 1 second 21 ms
30°
40 seconds 6 ms
40°
42 seconds 60 ms
50°
minute 22 seconds 45 ms
60°
minute 30 seconds
Conclusion
Several points can be drawn from the results of this experiment. The first is that the results are very similar to that of my prediction. It clearly shows an optimum temperature at which the catalyse will work, with an increase/decrease either side. I was surprised at the way the rate of reaction slowed above the optimum temperature. I had expected there to be a very sharp decline as soon as the optimum temperature was surpassed. This was the case, but the rate of decline slowed at one point, which contradicts my research. This may be due to an inaccuracy in my experiment, as my research suggests that enzymes fail to work after becoming denatured.
Another thing that can be learned from the results is that the speed at which the rate of reaction increases as the temperature rises seems to become greater. Between 20° and 30°, the increase is double the original rate of reaction. However, the increase in the rate of reaction seems to slow again between 30° and 40°. This could be for several reasons; that this is exactly what is supposed to happen and the rate of reaction does slow again when close to the optimum temperature, or that the optimum temperature lies somewhere between 30° and 40° and that the rate of reaction has reached its optimum temperature and is slowing again by the time the temperature reaches 40°. In any case, I would have to investigate this further in order to reach a firm conclusion as to the reason my results appear to show slowing of the increase in the rate of reaction between these temperatures.
I believe these results can be explained in the same way as my prediction, as the prediction was on the whole correct. Because the particles move faster as heat is applied, they bind with the enzymes quicker and more often so the rate of reaction speeds up. When the optimum temperature is surpassed, the enzymes begin to denature and cease to function, causing the rate of reaction to slow. The reaction will never stop completely, as hydrogen peroxide will break down naturally, even with no working enzymes present.
Evaluation
The data obtained in this experiment supports my conclusion well, although there were some results and trends that I couldn't explain. This may have been due to inaccuracies in the way the experiment was performed, or that I need to further my knowledge in order to explain the results.
I am convinced that the results show there is a correlation between the increase in temperature and the increased rate of reaction. This correlation would have been easier to work out had my measurements been slightly more accurate. The accuracy of these measurements could be improved by the use of a burette instead of a measuring cylinder, as it is a more precise piece of equipment and there are fewer margins for error. Another source of error may have been in the water baths, as they were supposed to be set at fixed temperatures to heat up the substances. One had to be very careful that the substances did not exceed their planned temperatures of there was danger of denaturing.
The experiment on the whole was a success, but it could be improved by the use of more accurate equipment and better organisation. Several assumptions had to be made in this experiment. When such assumptions are made, further work needs to be carried out to check these assumptions. I had to assume that all enzymes worked in the same way, but further work could be done with different enzymes and reactions to check this. Through the experiment, I measured the rate of reaction at 10° intervals. I think that further work should be done between 30° and 40° in an attempt to find the exact optimum temperature for the enzyme catalyse. Work between these temperatures would allow me to plot an accurate graph and explain the apparent slowing in the increase of the rate of reaction between these two temperatures in my current results.
GCSE Biology Coursework
Luke Hopwood 11B
