Worksheet for Morgan/Carter Laboratory #4 Enzymes

Ex. 4-1: EXPERIMENTAL METHOD AND THE ACTION OF CATECHOL OXIDASE Formulate a question about catechol oxidase and potato extract.

Propose a testable hypothesis for the presence or absence of catechol oxidase in potato extract.

Predict the result of this experiment (if/then).

What is the purpose of each of the three test tubes?

Which is the control tube?

Is more than one control tube necessary?

Design a table below in which you will record your results:

Explain your results in terms of your hypothesis.

Ex. 4-2: INHIBITING THE ACTION OF CATECHOL OXIDASE Propose a question about the activity of PTU.

Hypothesize about the nature of inhibition by PTU.

Predict the results of the experiment based on your hypothesis (if/then).

Which test tube is the control?

Why was the concentration of the catechol increased in test tube 2?

Why should the catechol be added to the test tubes last?

Design a table and record your results below:

Explain your results in terms of your hypothesis.

One member of your team is not convinced you have adequately tested your hypothesis. How could you expand this experiment to provide additional evidence to strengthen your conclusion?

Ex. 4-3: INFLUENCE OF CONCENTRATION, pH, AND TEMPERATURE ON THE ACTIVITY OF AMYLASE Experiment A: The Influence of Enzyme Concentration on the Rate of Starch Digestion Formulate a question about enzyme concentration and reaction rate.

Hypothesize about the effect of changing enzyme concentration on the rate of reaction.

Predict the results of the experiment based on your hypothesis (if/then).

Time of starch disappearance in different concentrations of the enzyme amylase: Tube 1 2 3 4 5 % Amylase Time of Starch Disappearance (sec)

What is the independent variable? Which axis is appropriate for this variable?

What is the dependent variable? Which axis is appropriate for this variable?

Explain your results in terms of your hypothesis. Describe the shape of the reaction rate curve as substrate concentration increases. What factors are responsible for the change in reaction rate?

Speculate about the shape of a curve measuring reaction rate if you had increased the concentration of enzyme, but held the concentration of substrate constant.

Experiment B: The Effect of pH on Amylase Activity Formulate a question about pH and reaction rate.

Hypothesize about the rate of amylase activity at various pHs.

Predict the results of the experiment based on your hypothesis (if/then).

Time of starch disappearance in different pH environments for the enzyme amylase: Tube 1 2 3 4 5 6 pH 4 5 6 7 8 9 Time of Starch Disappearance (min)

What is the independent variable? Which axis is appropriate for this variable?

What is the dependent variable? Which axis is appropriate for this variable?

Explain your results in terms of your hypothesis. Describe the shape of the reaction rate curve obtained with change in pH. What factors are responsible for the shape of this curve?

Experiment C: The Effect of Temperature on Amylase Activity Formulate a question about temperature and reaction rate.

Hypothesize about the rate of amylase activity at various temperatures.

Predict the results of the experiment based on your hypothesis (if/then).

Time of starch disappearance in different temperatures for the enzyme amylase: Tube 1 2 3 4 Temp. 80oC 37oC 22oC 4oC Time of Starch Disappearance (min)

What is the independent variable? Which axis is appropriate for this variable?

What is the dependent variable? Which axis is appropriate for this variable?

Explain your results in terms of your hypothesis. Describe the shape of the reaction rate curve as temperatures increase. What factors are responsible for the shape of this curve?

What do you think would happen to reaction rate in the tube incubated in ice if this tube, with enzyme and substrate already mixed, were placed in the 37oC water bath? (explain your answer in terms of the effect of various temperatures on enzyme structure and the rate of enzyme activity)

REVIEWING YOUR KNOWLEDGE catalyst cofactor

enzyme

coenzyme

substrate

competitive inhibitor

active site

non-competitive inhibitor

Compare and contrast competitive and noncompetitive inhibition.

Why does adding additional substrate overcome competitive but not noncompetitive inhibition?

Review changes in reaction rate as substrate concentration, pH, and temperature change. Explain the conditions or factors that are responsible for these changes.