Biology and Health Sciences

To be able to explain the role of enzymes in living organisms and how they are affected by temperature.

Learning objectives

After studying this unit, I should be able to:

Introduction

Look at the photographs below. Can you identify what is in the photographs? What is the relationship between them?

Try washing off the stain in a piece of cloth with water only, then using the detergent. Where was it easy to remove the stain?

Detergents which are used to remove stains act as enzymes. During the washing process, water and dirt are quickly combined together when the detergent is available to form a new product. This new product is easily removed from the cloth as we wash. On the other hand, water without detergent or soap  can take a long time to remove the stains.

Most metabolic reactions in the body are catalysed by enzymes; hence increasing efficiency of these processes in living organisms. It is important therefore to understand the nature, properties of enzymes and how they function.

Enzymes are biological catalysts that are protein in nature. They speed up or slow down the rate of chemical reactions in the body without being used up themselves.

6.1 Types of enzymes

Some enzymes are used within the cells that produce them. These are called intracellular enzymes, for example, respiratory enzymes and catalyse enzyme in the liver cells.

Other enzymes are transported from the cells that produce them to be used in other cells. These are called extracellular enzymes. Some examples of extracellular enzymes are those secreted into the alimentary canal to help with digestion.

There are also different types of enzymes based on the foods they act on, for example:

Activity 6.1: To investigate the presence of enzymes in living tissues

Requirements

Procedure

1. Label 4 test tubes as A, B, C and D.

2.  Measure 2cm3 of hydrogen peroxide solution and pour into each of the labelled test tubes.

3.  Into test tube A, do not add anything.

     • Note any changes or observations.

4.  Into test tube B, add some sand. Shake the test tube and note down any changes.

5.  Into test tube C add some pieces of potato that have been ground into a paste (using a pestle and mortar).

6. Repeat procedure (5) in test tube D using some liver which has been cut up then pounded in a mortar using a pestle.

7.  Record your observations in a table like the one below.

Living tissues contain an enzyme called catalase which breaks down hydrogen peroxide to water and oxygen.

The gas produced relights a glowing splint as seen in procedure 5 and 6. No gas is produced in Test tube 1 and 2 which do not contain living tissue.

6.2 Characteristics of enzymes

Discussion corner

1. Discuss with a classmate the characteristics of enzymes. Use the following questions as your guidelines:

2. Share your findings with other class members.

1.  Enzymes are protein in nature: all enzymes are made up of proteins.

2.  Enzymes are affected by temperature. They work best at specific temperatures; for example, enzymes found in human bodies work best at 37oC. This is called the optimum temperature.

• Very low temperatures inactivate enzymes. Therefore enzymes are not able to catalyse reactions.

• High temperatures beyond the optimum temperature denature enzymes. The structure of the protein molecule is destroyed by heat.

3.  Enzymes work best at specific pH. Different enzymes have a given specific pH at which they act best.

This pH is called optimum pH. Some enzymes work best at low pH (acidic medium) while others work best at high pH (alkaline medium). Most enzymes in the human body for instance work best at neutral or slightly alkaline pH. Examples are: lipases, peptidases and amylase. A few enzymes like pepsin that digests proteins in the stomach works best at an acidic pH of 2.

4. Enzymes remain unchanged after catalysing a reaction. Enzymes are catalysts and can therefore be used over and over again in small amounts without being changed.

5.  Enzymes catalyse reversible reactions. This means that they can change a substrate into products and the products back to the original substrate.

6.  Enzymes are substrate-specific. This means that an enzyme can only catalyse one reaction involving a particular substrate. This is because they have active sites which can only fit to a particular substrate whose shape complements the active site. For example, pepsin works on proteins but not on fats or starch.

7.  Enzymes work rapidly. Enzymes work very fast in converting substrates into products. The fastest known enzyme is catalase, which is found in both animal and plant tissues.

8.  Enzymes are efficient. This is best described by the fact that:

Activity 6.2: To investigate the effect of amylase on starch

Requirements

Procedure

1.  Put 2 cm³ of starch solution in a test tube and add 5 cm³ of 1% amylase solution to it. Label it A.

2.  In the other test tube, put 2 cm3 of distilled water and add 5 cm3 of 1% amylase solution. Label it B.

3.  Put the two test tubes in a water bath at 35oC and leave the set up undisturbed for 20 minutes.

4. Carry out a starch test on contents of test tube A and B.

5.  Record your observations in a table.

6.  Account for your observations.

7.  Share your work with the rest of the class.

Self-evaluation Test 6.1

1.  Distinguish between enzymes and other catalysts.

2. Explain the meaning of the following terms with reference to enzymes:

3.  The reaction below illustrates an enzyme catalysed reaction.

     State two characteristics of enzymes illustrated in the reaction.

4.  Explain why enzymes are said to be substrate specific.

6.3 Factors that affect enzyme activity

As mentioned earlier, enzymes are protein in nature and are very sensitive. Any change in the structure of an enzyme interferes with its ability to catalyse  a reaction. We therefore need to investigate the factors that affect enzyme activity. They include: temperature, enzyme concentration, substrate concentration and presence of inhibitor.

a) pH

Activity 6.3: To investigate the effect of pH on action of salivary amylase on starch

Requirements

Procedure

1.  Take a sample of starch solution and test for the presence of starch, and reducing sugars. Divide the remaining solution into three test tubes and label them as A, B and C.

2.  To test tube A, add 1cm3 of dilute hydrochloric acid and shake. Add 1cm3 of the enzyme amylase and shake.

3. To test tube B, add 1cm3 of dilute sodium hydroxide solution then 1cm3 of the enzyme amylase and shake.

4. To test tube C, add 1cm3 of distilled water into the test tube then add 1cm3 of the enzyme amylase and shake.

5.  Place the labelled test tubes into a water bath maintained at 37ºC for 20 minutes.

6.  Test the contents of the test tubes for the presence of starch and reducing sugars.

7.  Record your results and conclusions in a table like table 6.2.

8.  Discuss your findings with a classmate.

9.  Share your findings with the rest of the class.

10. Account for your results in the experiment above.

Different enzymes have different pH at which they work best. Most enzymes in the human body however, work best at pH 7 (neutral pH). Some digestive enzymes have different optimum pH though. For instance pepsin digests proteins in the stomach at a pH of 2 while trypsin digests proteins in the duodenum at a pH of 9.  Extreme pH causes enzymes to denature and permanently lose their function.

b) Temperature

Activity 6.4: To investigate the effect of temperature on action of ptyalin

Requirements

Procedure 

1.  Place 2 cm³ each of starch solution into three different test tubes labelled A, B and C.

2.  To each test tube, add 1cm3 of amylase enzyme.

3.  Immerse test tube A into a beaker of cold water (preferably with icecubes).

4.  Put test tube B in a water bath maintained at 37ºC.

5.  Boil the contents of test tube C.

6.  Test the contents of each test tube for the presence of starch and reducing sugars.

7.  Record your results in a table like table 6.3.

Enzyme activity is higher at high temperature since molecules gain more kinetic energy. There is therefore a higher chance of substrates bumping into enzyme’s active site. The temperature at which enzymes work best is called optimum temperature.  Temperatures lower than the optimum cause enzymes to be inactive. Therefore they work at a slow rate.

Higher temperatures than the optimum temperature destroy enzymes. This is because such temperatures alter or change the structure of the enzyme. This destroys the binding site; enzymes are therefore said to be denatured. A denatured enzyme cannot function. Most enzymes begin to denature at 40^oC. However, enzymes of bacteria found in hot springs may begin to denature at temperatures higher than 80^o C.

Note: Other factors of that affect enzyme activity are: substrate concentration, enzyme concentration, presence of inhibitors and co-factors.

Self-evaluation Test 6.2

1.  Which of the following has an active site?

2. Name two factors that destroy the shape of the active site of an enzyme.

3.  Enzyme pepsin is a protease that digests proteins in the stomach.

4. An enzyme Z was extracted from the alimentary canal of a certain mammal. The effect of pH on the activity of this enzyme was investigated at 38o C. The results are shown in the figure below.

6.4  Mode of enzyme action

Activity 6.5: To investigate the fitting of substrate to enzyme active site

Requirements

Procedure

• Working in groups, each member of the group should try to find the right key for the two padlocks.

Study questions

(a) Were you able to find out the right key?

(b) What conclusion can you make from the activity?

(c) What do the following correspond to with regard to enzymes and substrates?

      •    Key                   •   Padlock hole

Enzymes are very specific in nature. This means that they can only catalyse a specific reaction. The shape of the substrate must complement the shape of the enzyme’s active site. This enables fitting of the substrate into the enzyme active site and enables the substrate to be changed into products. The 'groove' in an enzyme where the substrate fits during the reaction is called active site. When the substrate fits into the active site, an enzyme-substrate complex is formed. This reaction complex can be explained by a key and lock mechanism as illustrated below.

Further Activity 6.6:  To investigate the action of enzyme amylase

Requirements

Procedure

1.  Label three different petri dishes as P1, P2 and P3

2. Put the different concentrations of amylase in the three different Petri-dishes.

3.  Put a small disc of filter paper in each of the amylase solutions in the petri-dishes. 3.  Cut a circular piece from the plain photocopy paper that can fit at the bottom of a petri-dish.

4.  Place the circular paper at the bottom of an empty petri-dish and irrigate it with iodine solution.

5.  Pour away excess iodine solution.

6.  Using the teat pipette, pour water on the circular paper and rinse it.

7. Using the blunt forceps, remove the small discs of filter paper from the respective solutions and place them on the stained piece of circular paper in the petri-dish.

8.  Cover the petri-dish and leave the set up for 15 minutes.

9.  Using the forceps, gently lift the circular pieces of paper from the petri-dish one at a time taking care not to tear the paper beneath.

10.  Observe the circular portions where you removed the pieces of filter papers.

11.  Record your observations in a table.

Study questions

(a) Explain the observations you have recorded for the stained piece of paper where the small disc of filter paper was placed.

(b) Suggest three ways you could improve this investigation.

Self-evaluation Test 6.3

1.  Amylase is a digestive enzyme found in the saliva. Which of the following food substances does it breakdown?

2. What does the key and lock hypothesis refer to with regard to enzyme action?

3.  Draw a graph that shows the effects of increasing temperature in an enzyme catalysed reaction.

Unit summary

• An enzyme is a substance produced by a living organism that acts as a catalyst to bring about a specific biochemical reaction.

• A catalyst is a substance that increases the rate of a chemical reaction without itself undergoing any permanent chemical change.

• Enzymes are characterized by their specificity, sensitivity to temperature and pH change, and working very fast.

• Factors affecting enzyme activity include pH and temperature range.

• The substrate must fit into the active site of an enzyme for a reaction to take place. This forms the key and lock mechanism.