Cell metabolism and Enzymes

Metabolism (all of the chemical reactions in a living thing) Anabolic reactions Catabolic reactions Small molecules are combined to make larger molecules. Energy is needed Large molecules are broken down into smaller molecules. Energy is given out.

Sources of energy in cells for anabolic reactions 1.Solar energy Sunlight is a source of energy capable of being absorbed by cellular pigments e.g. chlorophyll. 2. Cellular energy Catabolic reactions in cells release energy

Enzymes

Enzymes – the key to it all Almost all chemical reactions that take place inside living things are controlled by enzymes. Enzymes are biological catalysts – they change the rate of reactions without getting used up themselves. Every enzyme is a protein and has a specific 3D shape.

Enzyme Action The substance that an enzyme acts on is its substrate The substance(s) that the enzyme forms is called the product(s)

Catabolic enzyme Catalyse is an enzyme that converts hydrogen peroxide into water and oxygen Catabolic = breaks a big molecule into smaller ones

Anabolic Enzyme DNA Polymerase is an enzyme that builds DNA nucleotides from tiny nucleotide units in our cells Anabolic = makes small molecules into bigger ones

Consider This Amylase is an enzyme found in saliva. It breaks starch molecules into smaller maltose molecules What is the substrate? What is the product? Will amylase break fats into fatty acids + glycerol? Why? Is this an example of a catabolic or an anabolic reaction? Why?

And This DNA Polymerase is an enzyme that builds DNA using tiny molecules in our cells What is the substrate? What is the product? Do you think DNA polymerase could do the reverse reaction? Why?

Learning Check What is meant by a substrate? What is the unit formed when an enzyme combines with its substrate called? Give an example of an anabolic reaction Give an example of a catabolic reaction

Enzymes and Temperature

Mandatory experiment Investigation 1 How does temperature effect the rate of reaction of an enzyme?

1.5g of chopped up celery 2.1 drop of washing up liquid 3.10mls of Buffer solution ALL INTO A GRADUATED CYLINDER Step A

1.5ml of Hydorgen peroxide 2.The graduated cylinder and contents ALL IN A WATER BATH UNTILL THEY ARE THE SAME TEMPERATURE AS THE WATER BATH Step B

Add the hydrogen peroxide to the graduated cylinder. Time for 1 minute and see the volume of oxygen made Step C

Temperature and enzymes At very low temperatures molecules are moving slowly and enzymes work slower because of less collisions with substrate As temperature increases molecules start to move and bump into each other causing the rate of reaction to increase

Temperature and enzymes Above certain temperatures enzymes start to lose their shape the rate of reaction falls. When the shape is fully lost the enzyme is said to be denatured this is usually a permanent condition Human enzymes work best at body temperature (37°C) Plant Enzymes work best at °C Rate of reaction Temp ( o C) 37

Check your learning Why does temperature have an effect on the rate of reaction? Why do enzymes not work well at low temperatures? As temperature increases what happens to enzyme activity? What is the optimum temp for enzymes in plants and humans? What happens when an enzyme loses its shape?

Enzymes and pH

Mandatory experiment Investigation 2 - To investigate the effect of pH on the rate of enzyme action

pH pH scale goes from Less than 7 is acidic 7 is neutral More than 7 is basic (or alkaline)

To investigate the effect of pH on catalase activity

Step 1 Finely chop the celery. Weigh 5 g of the chopped celery.

Step 2 Add 10 ml of your pH buffer 4, one drop of washing-up liquid and 5 g of celery into a graduated cylinder.

Step 3 Add 5 ml of hydrogen peroxide to a boiling tube.

Step 4 Place the graduated cylinder and the boiling tube in a water bath at 25 °C – until they reach the required temperature

Step 5 Add the hydrogen peroxide into the graduated cylinder and record the volume immediately.

Step 6 Time for 2 minutes and record the final volume.

Enzymes and pH pH is important for enzymes as it can make enzymes change their shape! The best ph (optimum pH) is usually pH 7 Outside this they lose their shape and get denatured Rate of reaction 0 pH 14

Check your learning.. What is an enzyme? Why are they needed for our metabolism? What does rate of reaction of an enzyme mean? What does an enzyme act on? What effect does temperature have on the rate of activity of an enzyme? What effect does pH have on the rate of activity of an enzyme?

Higher level – The active site theory of enzymes

How enzymes work Enzymes are not flat they have a 3D structure The active site is the specific part of the enzyme which combines with the substrate.

Enzymes – active site theory Because of the special shape of the active site enzymes are very specific about the reactions they catalyse. If the shape of the active site changes then the enzyme cannot work and is denatured.

enzyme reactant active site 1. Substrate fits into the active site 2. Enzyme- substrate complex is formed. The enzyme changes the structure of the substrate products 3. Products leave the active site The enzyme changes back to original shape

What conditions could denature an enzyme?

Mandatory experiment Investigation 4: the effect of heat denaturation of the activity of an enzyme

Put 5g of chopped celery into two boiling tubes in water baths at 100 O C and 20 O C for 10 minutes. Remove and cool.

Make up 2 graduated cylinders with.. 10cm3 of pH buffer 9 1 drop of washing up liquid Add boiled celery to A Add unboiled celery to B

Step 3 In a water bath at 20 o C add 5 cm 3 of hydrogen peroxide to both graduated cylinders Measure the volume of oxygen gas produced in two minutes

Check your learning… What is meant by the specificity of an enzyme? How can the Active Site Theory explain the specificity of enzymes? How can an enzyme be denatured? Give two features of a denatured enzyme?

Immobilising enzymes

Bioprocessing Bio-processing is the use of enzyme controlled reactions to produce a product Traditionally micro- organisms such as bacteria and yeast were used but since the 1950’s enzymes are being used

Immobilising enzymes Enzymes are often immobilised or fixed (attached to each other or an inert substance) so that they can be used repeatedly in a bioreactor

How to immobilise enzymes The enzyme can be trapped in beads using sodium alginate and calcium chloride

Mandatory experiment Investigation 4: To prepare an immobilised enzyme and examine its application

Step 1 Making up the solutions

In one beaker – make up a sodium alginate solution

In a second beaker – make up the yeast solution

In a third beaker – make up calcium chloride solution

Step 2 Trapping yeast enzyme sucrase in beads

Mix the yeast with the alginate solution

Pull the yeast/alginate solution up into the syringe

Drop the solution from the syringe into the calcium chloride solution

Step 3 Investigating the activity of the immobilised enzyme compared to the normal enzyme

Filter the hardened beads through a sieve and rinse with water.

Fill up seperating funnels with free yeast and immobilised yeast

Dissolve 1 g of sucrose in 100 ml of distilled water. Pour 50 ml into each separating funnel.

Immediately test the products in the beakers with glucose strips.

Repeat test every 2 minutes until glucose appears in both beakers.

Run off the remaining product from each funnel into the beakers and compare the turbidity * of the solution from both funnels.

Advantages of Immobilised Enzymes 1.A gentle procedure is used so the efficiency of enzyme is not affected 2.Immobilised enzymes can be easily recovered from the product so you can get a pure sample of product easily 3.Immobilised enzymes can be reused this cuts costs

Uses of Immobilised Enzymes Immobilised sucrase converts sucrose into glucose to fructose which is used to make sweet foods like marshmallow

Learning check What is an immobilised enzyme? How are enzymes immobilised? What are the advantages of immobilising enzymes? Can you give an example of what immobilised enzymes are used for?