Problem: To determine the effect of substance X on activity of a mammalian enzyme, an assay is performed in which the initial concentration of the enzyme's substrate [S] is varied and the initial rate of reaction (v) is measured in the presence or absence of X. The same amount of enzyme is used in all reactions and the reactions are incubated under identical conditions so that any effects can be attributed to the treatment being investigated. The results of this experiment are presented in the table.
|
Without X |
With X |
| Concentration of substrate, [S] |
Rate of reaction, v |
Rate of reaction, v |
| 2.5 |
10 |
7.5 |
| 5 |
23 |
13.2 |
| 10 |
30 |
20.5 |
| 20 |
36 |
28.1 |
| 40 |
40 |
34.6 |
| 80 |
42.4 |
39.1 |
| 120 |
44 |
43 |
1) Identify one condition/factor (apart from the concentration of the enzyme) that would be particularly important to keep constant in such an experiment. For the condition that you have identified, explain why it is important that it is kept constant.
b) A Michaelis-Menten plot is a plot of the initial rate of an enzyme-catalysed reaction, v against concentration of substrate at the start of the reaction, [S] . Use the data in Table 2 to draw a single graph showing the Michaelis-Menten plots for the enzyme in both the absence and in the presence of substance X.
Include a figure legend for your graph (a title is not required). The identity of the two plots should be made clear either through the inclusion of an appropriate key or explanation in the legend.
c) From your graph, estimate the values of v^max and KM for the enzyme in the absence and in the presence of X, remembering to include the appropriate units.
d) What can you conclude about substance X? Justify your answer with reference to vmax and KM values and explain the effects observed in the experiment.