Temperature Effects on Enzymes

Kevin Rivera
kjr0107@unt.edu

Bio1730.526
October 7, 2014

Schmidt
Wednesday 8PM-11PM

Abstract
This experiment examined how heat can affect an enzymes function. By using a combination of
a liquid with a PH of 7, Iodine, unheated enzymes, and heated enzymes, it was clear to see that
the heated enzyme denatured. Denaturation is a loss of tertiary (and often secondary) protein
structure not involving covalent bond cleavage. (Dines et al., 1996, p. 1) Therefore when an
enzyme denatures, it unfolds and loses its catalytic ability. Denaturation was evident when the
heated enzyme started to lose its ability to change the substrates color very quickly as time went
on. Therefore it is incontrovertible that heating an enzyme causes it to denature and in turn
causes the enzyme to lose its function; heating an enzyme causes loss of function.

Introduction
An enzyme is a catalyst, which means it will accelerate the rate for something that is already
taking place. If an enzyme ceases to function, it can no longer catalyze a reaction, which will kill
a cell. In this experiment we had our experimental group A and our control group B as two
separate rows on a porcelain spot plate. For both groups we used a liquid with a PH of 7 and
Iodine. However, for our experimental group we used heated enzymes, and for our control group
we used enzymes at room temperature, roughly 72 degrees Fahrenheit. Over time the heated
enzymes were unable to change the color of the substrate, whereas the unheated enzymes could
clearly change the substrates color. Therefore, temperature affects color change directly. If a
color change occurs when an enzyme is at an optimal temperature then heating an enzyme
should slow down or even halt the color change of a substrate due to denaturation.

Materials and Methods

The following materials are required: two test tubes, one porcelain spot plate, boiling water, two
pasteur pipettes, toothpicks, substance with a pH of 7, enzyme, starch, and Iodine. When
referring to the porcelain spot plate use the diagram below:

Add 2mL of the enzyme to two test tubes and put one of them in boiling water for 5 minutes. On
the porcelain spot plate, in the first two rows from the top, place five drops of starch on every
well. Then place 2 drops of the pH7 buffer in each well that already contains starch and mix
these two substances thoroughly with a toothpick. As close in time as possible, place one drop of
the unheated enzyme on every well in the top row and one drop of the heated enzyme on every
well in the middle row. Right after that add 1 drop of Iodine to each well in column 1 and stir. At
5-minute intervals add 1 drop of Iodine to each column and record your observations. When you
are finished wash all lab equipment and carefully dispose of the substances in the wells in a BioHazard bag.

Results
Over time, the heated enzyme was incapable of changing the substrates color, while the
unheated enzyme clearly changed the color of the substrate. In the first column there was no

difference between the heated and unheated enzymes because no time had elapsed. After 5
minutes the second column had a very slight difference between the heated and unheated
enzymes since very little time had elapsed. However, in the third and fourth columns there is a
very clear difference between the heated and unheated enzymes since there is a noticeable color
difference. The only difference between multiple groups that also did the experiment is the
intensity of the color difference. Below is a picture of the results:

The hypothesis on the introduction is accepted since when the enzyme was heated there was no
color change because denaturation occurred.

Discussion
The heated enzyme had no color change because it denatured. The unheated enzyme definitely
had a color change since it did not denature. The color difference in the unheated and heated
enzymes was expected. However, the experiment was not perfect because there was
contamination in either the heated enzyme or Iodine. The color change was not even for the
heated enzyme. The color change should be svn because of the 5 minute intervals. We also did
not mix the final solutions in each well for the exact same amount of time. Perhaps a machine of
some sort could be used to make all mixing perfectly even for the same amount of time. By this

point it is incontrovertible that heating an enzyme will cause no color change to the substrate in
this experiment but what would happen if we refrigerate the enzyme?

Conclusion
Denaturation of an enzyme will cause it to lose its catalytic ability. In this case that was shown
by the substrates lack of color change. Therefore, when an enzyme is heated it ceases to work.

References
Daniel, R., Dines, M., Petach, Helen. July 1st 1996. The denaturation and degradation of stable
enzymes at high temperatures. PMC Journal. Pages 1-11.

Daniel, R., Danson, M., Eisenthal, R., Lee, C., Peterson, M.