Practical Investigation: Protein Digestion by an Enzyme

PRACTICAL INVESTIGATION: PROTEIN DIGESTION BY AN ENZYME

Gelatine is a complex protein formed by the hydrolysis of collagen, which is found in animal cartilage and bones. It is extracted by boiling the tissue in water.

When dissolved in water, gelatine sets like a jelly. Gelatine is used in foods, photographic products and adhesives.

Bromolase is a naturally occurring enzyme found in fresh pineapple juice. Bromolase digests gelatine and prevents it from setting.

EXPERIMENT 1

Aim: To investigate the digestion of gelatine using the enzyme Bromolase

Hypothesis:

Method:

1.  Make up a 10% weight/volume solution of gelatine (50g to 500mL)

2.  Label test tubes A1, B1, C1, A2, B2, C2 and place 5mL of the gelatine solution in each

3.  Mix 5mL of water into test tube A1 and A2

4.  Mix 5mL of fresh pineapple juice into tube B1 and B2

5.  Mix 5mL of boiled pineapple juice into tube C1 and C2

6.  Incubate the tubes at 40oC for 15 minutes

7.  Transfer the tubes to an ice bath

8.  Check the tubes for solution setting using the 0, +, ++ indicators (0 = not set, + = partially set, ++ = fully set) and record in the table below.

9.  Recheck the tubes every 3 minutes for 30 minutes.

Results:

Conclusion:

Discussion:

EXPERIMENT 2

Aim: To investigate the effect of changing concentration of Bromolase on enzyme activity

Hypothesis:

Method:

1.  Make up gelatine solution as in experiment 1

2.  By adding water to pineapple juice make up 2 series of dilutions of the juice. The dilutions are 100%, 80%, 60%, 40%, 20%, 0% (straight water)

3.  Place 5mL of each dilution in a separate labelled test tube

4.  Add 5mL of gelatine to each tube

5.  Incubate the test tubes for 15 minutes at 40oC

6.  Transfer the tubes to an ice bath

7.  Vheck the tubes for the solution setting using the 0, +, ++ indicators described in experiment 1 and record in the table below

8.  recheck the tubes every 3 minutes for 30 minutes

Results:

Draw a graph of this data

Conclusion:

Discussion:

EXPERIMENT 3

Aim: To investigate the effect of increasing temperature on the activity of Bromolase enzyme

Hypothesis:

Method:

1.  Make up the gelatine solution as in experiment 1

2.  Put 5mL of pineapple juice into 12 labelled test tubes

3.  Prepare 6 beakers at different temperatures to act as water baths. The temperatures should range from 0oC to 80oC

4.  Place a 2 tubes of juice into each of the separate water baths

5.  When the juice temperatures have reached the temperatures of the water baths, add 5mL of gelatine to each tube

6.  Incubate the tubes for 15 minutes in the water baths

7.  Transfer the tubes to an ice bath

8.  Check the test tubes for the solution setting using the 0, +, ++ indicators described in experiment 1 and record in the table below

9.  Recheck the tubes every 3 minutes for 30 minutes

Results:

Draw a graph of these results

Conclusion:

Discussion:

EXPERIMENT 4

Aim: To investigate the effect of pH on the activity of Bromolase enzyme

Hypothesis:

Method:

1.  make up the gelatine solution as in experiment 1

2.  Put 5mL of pineapple juice into 6 labelled test tubes

3.  Add 1mL HCl into two labelled test tubes

4.  Add 1mL NaOH into the another 2 labelled test tubes

5.  Add 1mL distilled water into the final 2 test tubes

6.  Add 5mL of gelatine to each tube

7.  Incubate the tubes to 15 minutes at 40oC

8.  Use a pH meter to check the pH of each solution

9.  Transfer the tubes to an ice bath

10.  Check the tubes for the solution setting using the 0, +, ++ indicators described in experiment 1 and record in the table below.

11.  Recheck the tubes every 3 minutes for 30 minutes

Results:

Conclusion:

Discussion