Investigating The Effect of Temperature on the Structure of an Enzyme

Investigating The Effect of Temperature on the Structure of an Enzyme
Introduction:For my gcse Biology assessment I will be investigating the enzymeamylase with the substrate starch. This reaction, which I am going toinvestigate, is called the protein test for starch.Aim:My intention for this observation is to examine how the enzymecatalyses are affected by changes in temperature.Safety Precautions:In this investigation I am going to make sure that everything is assafe as possible and prevent any accidents from occurring.1) I will wear safety glasses and a lab coat to prevent avoidsdanger.2) I will tie my hair back, which will prevent any accidents fromoccurring.3) I will keep my area around me clear to prevent any accidents fromoccurring.4) Benches should be tucked under the table so that nobody tripsover
5) As Iodine indicates the symbol means irritant (harmful). Therefore I will be careful with this chemical that it doesn?t get to my skin. 6) Chemicals should be placed in the tray so that if they spill they don?t drop all over the place and accidents from occurring could be stopped. Apparatus: The apparatus that I require for this investigation is: 1) Test tubes (4): These will be used to place the amylase and starch solution in 2) Test tube rack: This will be used to place the test tubes into 3) Graduated pipettes (2): These will be used to measure the amount of Amylase/starch solution needed. 4) Glass rods (2): These will be used to place the mixture of the amylase and starch solution into the spotting tiles. 5) Water baths maintained at 100C, 200C, 300C, 400C, 500C and 600C (6): These will be used to heat up the solutions. 6) Thermometers (2): These will be used to tale the temperature reading of the solutions. 7) Backers (2): These will be used to dip the glass rods in each time that they are used for the solution. 8) Amylase solution 1%: This will be used as an enzyme for this reaction. I will use 1cm3 of this solution. 9) Starch solution 1%: This will be used as a substrate for this reaction. I will use 1cm3 of this solution. 10)Iodine solution: This will be used to test the amylase and starch solution. 11)Spotting tiles (2): These will be used to test the amylase and starch solution with iodine. 12)Stop Watch (2): These will be used to take the time of how long it takes the enzyme to catalyse. Scientific Knowledge with prediction: Within all cells many chemical reactions are occurring. Without presence of enzymes in a biological system the reactions that occur will either be extremely slow or the reactions might not occur at all. Enzymes are there to increase the rate at which the reaction works. What are enzymes? An enzyme is a globular protein that acts as a catalyst for a chemical reaction. Enzymes are known as biochemical catalysts that speeds up the reaction. If we would imagine the reaction without enzymes occurring, then the reaction would take place extremely slowly. The substance, which is there in the reaction at the beginning, is called the substrate. The enzymes however are very specific. They will only join with one type of substrate. There are two types of enzymes: 1. Intracellular enzymes: These enzymes are the reactions, which occur inside the cells. 2. Extra cellular enzymes: These enzymes are the reactions, which occur outside the cells. [image] How do enzymes work? Enzymes work by combining with the substrate, which is present at the beginning of the reaction. The substrate will fit into the active site of the enzyme, although the enzymes and substrate are very specific as only certain substrate molecules will fit into the enzymes active site. This theory is called the lock and key theory. In the reaction as the substrate molecules fit into the enzymes, an enzymes/substrate complex is formed. The new substrate at the end of the reaction gives out products from the reaction. Therefore the word equation for this would be: [image]Amylase + Starch Maltose Heating the enzymes will cause more collisions between enzymes and substrates. This is because all the molecules in the solution will move around quicker, therefore the enzymes and the substrate will collide with each other more regularly and there will be more kinetic energy in the molecules. I am aware of the fact that enzymes are affected by temperature, although the temperature will need to be at its optimum level. Being optimum is the condition at which enzymes work best, this is around 370C-400C. Consequently this means that only till a certain temperature the enzyme will speed up its rate of reaction. As a result the enzymes, which are over the optimum temperature, they will be denatured, this is not a reversible reaction. Being denatured means that the active site in the enzyme will change its shape so that the substrate molecules will no longer be able to react with the enzyme. Using the knowledge that I have gathered I will now predict that: As the temperature increases till its optimum the rate at which the reaction occurs will also increase. From my research I would therefore say that the optimum temperature is about 370C-400C, so if the temperature reaches higher it will therefore be denatured. Also I have found out that the Q10 law says that: For every 100C raise in temperature the rate of reaction will double, up to about 400C when the enzymes will start to denature. -????????????????????????????????? Anther condition that could affect the reaction rate of the enzymes and substrate is the pH of the solution. The pH is to measure how acidic or alkali a solution is. Amylase has the optimum pH of 7-8 that tells us that this is a neutral to alkali solution. So having a higher pH means that the enzyme will start to lose its shape so in other words it is denatured. Anther condition, which could affect on the rate of reaction of the enzymes and substrate, is the enzyme concentration, but this is provided that the substrate concentration should be kept constant. As there are lots of enzyme molecules in the reaction, it will occur faster. The reason for this is because it?s more likely that the substrate molecules will bump into the active site of the enzymes more frequently. Anther condition that could affect the rate at which the reaction works is the concentration of the substrate. As long as there are enough enzyme molecules when the substrate concentration increases the substrate molecules will bump into the active site of the enzymes. Although if there are too many substrate molecules then they cannot react very fast as the molecules are queuing up for the active site of the enzymes. Before planning my experiment I have done a preliminary study, which has helped me to plan out my investigation, which helped me, as I knew how the reaction works so, how the affect of temperature will speed up the reaction. Also this gave me an idea of what the investigation is about. Method:
I will get my beaker of cold water and add some ice.
I will then take the reading of the water with a thermometer and
add some ice till it reaches 100C.
I will then opened the amylase solution bottle and fill it in the
graduated pipette of 1cm3.
Then from the graduated pipette I will release the amylase
solution in the test tube.
I will then do the same for the starch solution.
I will then place both test tubes in the water bath maintained 100C;
I will leave this for 3 minutes.
Whilst the solutions are in the water bath I will take the iodine
and put a drop into each tile in the spotting tile.
I will then after 3 minutes pour the starch solution in to the
amylase solution test tube and mix well.
I will place this test tube back into the water bath and start the
stopwatch.
Every 30 seconds I will take some of this solution with a glass
rod and mix it up with the iodine in one of the tile.
Each time I will wash the glass rod and wipe it with a kitchen
towel.
When the colour will change to yellowish brown I will record the
time it took.
I will repeat this 100C experiment anther 2 times so I can take an
average of the results.
I will then repeat this procedure for the temperature 200C, 300C, 400C, 500C and 600C. Results: Temperature (() C) Time (seconds) Average Time (Seconds) 1st Test 2nd Test 3rd Test 10 725 800 708 744 20 576 590 600 589 30 490 477 400 455 40 340 387 359 362 50 550 489 587 542 60 687 620 654 654 Conclusion: Temperature/Time graph: The temperature time graph shows me that when the temperature has increased the time it takes for the enzyme to catalyse will decrease. The significance behind this is that at a higher temperature the enzymes will catalyse faster. This is because the time it takes for the enzymes to catalyse under 400C is due to kinetic energy. Therefore the number of collisions between the substrate and the enzymes active site has increased. Although by looking at my graph I can say that the optimum temperature is 400C, the reason for this is because at 400C the enzymes are catalysing the fastest. However at 500C the time taken for the enzymes is longer, although the enzymes and substrate are not catalysing they are getting destroyed, so we say that they are denatured as the shape of the active site changes. Therefore the substrate molecules will no longer be able to fit into the active site.

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