Module Enzymes By Ms Cullen

What is an enzyme? Q: can you remember any enzymes that you encountered at GCSE? A:

What is an enzyme? Did you know ? Without enzymes the reactions in our bodies would be too slow to sustain us. An organisms metabolism consists of thousands off different reactions, all catalysed by different enzymes. The name of the substance that enzymes act on is called the substrate. Some enzymes are intercellular eg hydrolases in lysosomes, and ATPase in mitochondria. And others are extracellular eg digestive enzymes.

Intercellular enzymes Catalase is an example of an intercellular enzyme. It ensures hydrogen peroxide, a toxic product of many metabolic reactions is broken down into harmless oxygen and water. 2H 2 O 2 2H 2 O + O 2 This particularly occurs in hepatocytes, liver cells. They contain high concentrations of catalase. catalase

Extracellular enzymes Digestive enzymes are released from cells to break down large nutrients in form of polymers to smaller molecules. Amylase - Digests starch.

The Digestive System

Extracellular enzymes Protease - Digest Proteins.

How do enzymes work? The shape of the enzyme’s active site is complementary to the shape of the substrate molecule. The active site provides a site for the reaction to occur.

The lock-and-key hypothesis

The induced-fit hypothesis

Catabolic enzymes –

Anabolic enzymes –

Carry out Activity 27: ‘Effect of enzyme concentration on the rate of reaction’

Effect of enzyme concentration on the rate of an enzyme- controlled reaction

Activation Energy In many reactions the substrates could not be converted into products unless it was given some extra energy. This energy is known as the activation energy. One way we can do this is to give the reactants more kinetic energy by heating them. As our bodies work at 37˚C we can not increase the temperature to above this without damaging proteins and enzymes and some reactions need energy at higher temperatures. Enzymes are the answer to this problem as they decrease the activation energy of the reactants that they catalyse.

Activation Energy Enzymes decrease the activation energy by holding the substrate molecules in such a way that their molecules react more easily. This allows reactions to occur more rapidly and at lower temperatures.

Activation energy (maltose to glucose) ‏

How the enzyme maltase lowers the activation energy needed to convert maltose to glucose

Carry out Activity 26: ‘Effect of substrate concentration on catalase’

Effect of substrate concentration on the rate of an enzyme- controlled reaction Vmax

The effect of temperature on enzyme activity Q: How does temperature usually affect a chemical reaction? A:

The effect of temperature on enzyme activity This is true of reactions controlled by enzymes too at first. However if the temperature gets too high, usually above 40˚C, then the structure of the protein will begin to change. This will change the shape of the active site and prevent enzyme/substrate complexes forming. This is irreversible. The enzyme is said to be denatured.

The effect of temperature on the rate of an enzyme-controlled reaction Usually the rate of reaction doubles for every 10˚C rise until optimum temperature is reached.

Temperature Coefficient – Q 10 The change in the rate of a reaction for each 10˚C rise in temperature is called the temperature coefficient or Q 10. At suboptimal temperatures the Q 10 for enzyme-catalysed reactions is usually 2 (the rate doubles for each 10˚C rise in temperature). The rate continues to rise until it reaches a peak at the optimum temperature.

Calculating Q 10 Q 10 = rate of reaction at T + 10˚C rate of reaction at T (T is temperature)

Q 10 = rate of reaction at T + 10 ˚C rate of reaction at T Q 10 = rate of reaction at 30 ˚C (20 ˚C + 10 ˚C) rate of reaction at 20 ˚C = 3.5 = For this reaction the rate of reaction doubles for each 10˚C rise in temperature. Calculating Q 10

Carry out Activity 25: How temperature affects the rate of milk hydrolysis

The effect of pH on enzyme activity Most enzymes work best in neutral conditions, around pH 7. pH measures the amount of H + ions in a solution, the lower the pH the more hydrogen ions present. The H + ions can interact with the R group of the amino acid, changing the tertiary structure of the protein. H + ions can interfere with both hydrogen bonds and ionic bonds.

The effect of pH on enzyme activity If the pH conditions are very different to the optimum pH for an enzyme it will become denatured. The tertiary structure will change as mentioned. Also H + ions are attracted to negatively charged amino acids at the active site. This inteferes with the substrates binding ability.

The effect of pH on enzyme activity

Carry out Practical 4.2: To investigate the effect of varying pH on the rate of a reaction catalysed by trypsin

Optimum pH graph versus rate of reaction

Pepsin and trypsin – rate graph

Effect of enzyme concentration The number of substrate molecules an enzyme can turn into products in one minute is called the turnover number. Chymotrypsin – 6000 Catalase – Carbonic anhydrase – With unlimited substrate, optimum pH and optimum temperature, rate of reaction will be directly proportional to enzyme concentration.

Initial reaction rate comparisons a b Initial rate calculated as gradient of tangent a = rate in cm 3 per second b

Calculating Gradients on graphs Gradient = change in y change in x

Enzyme inhibitors A substance which slows down or stops an enzyme controlled reaction is known as an inhibitor. There are 2 types of inhibitor: 1.Competitive inhibitors 2.Non-competitive inhibitors Inhibitors that seriously disrupt enzyme-controlled reactions can act as metabolic poisons. The death cap fungi contains a toxin, alpha- amanitin, which inhibits the enzymes that catalyse the production of RNA from DNA. This leaves cells unable to produce proteins, which is deadly.

How a competitive inhibitor works

Effect of concentrations of inhibitor and substrate on the rate of an enzyme-controlled reaction

How non-competitive enzyme inhibitors work

Rate of an enzyme-controlled reaction with and without a non- competitive inhibitor

End-product inhibitors

Carry out Practical: The effect of an inhibitor on enzyme activity with bananas.

Cofactors A molecule or ion that helps an enzyme work. It may be a coenzyme, a prosthetic group or an activator (inorganic ion). Usually simpler molecules, compared to larger coenzyme molecules. They may change the shape of the active site to make it work more efficiently. Many cofactors are made from vitamins. An example is Cl - which is the cofactor for the enzyme amylase.

Enzymes at work: Cofactors; Coenzymes Small, organic, non-protein molecules. Bind with active site just before or at same time as the substrate. Take part in the reaction but can be recycled and used again. Coenzymes usually carry chemical groups from one enzyme-controlled reaction to another. Vitamins are a good source of coenzymes. Examples:

Enzymes at work: Cofactors; prosthetic groups Prosthetic groups are coenzymes that are permanently attached to an enzyme. They contribute to the shape, charge & properties of the enzyme. The enzyme carbonic anhydrase has Other proteins have these too. The prosthetic haem group in haemoglobin allows it to carry oxygen. Hb could not do this without it.

Enzymes at work: Cofactors; inorganic ion cofactors These may bind to substrate or enzyme and allow more effective binding to form the enzyme-substrate complex. They do this by changing the overall charge (they are ions) and sometimes the overall shape of the ES complex. Obtained from food/minerals/supplements. Examples: Mg 2+ is needed for the enzymes of protein synthesis to work. Amylase requires Cl - ions to work.

Enzymes at work: Cofactors; inorganic ion cofactors The key inorganic ions that are involved in biological processes are below. You need to learn these! Cations: Calcium ions (Ca 2+ ) Sodium ions (Na + ) Potassium ions (K + ) Hydrogen ions (H + ) Ammonium ions (NH 4+ ) Anions: Nitrate (NO 3 - ) Hydrogencarbonate (HCO 3 - ) chloride ions (Cl - ) Phosphate ions (PO 4 3- ) Hydroxide ions (OH - )

Precursor Activation Enzymes are often produced in inactive forms, called precursor enzymes, to prevent them causing damage to cells or tissues when they are released. To be activated the precursor enzyme needs to undergo a change in the tertiary structure, so that the active site can be activated. Usually the addition of a cofactor will cause this to happen.

Precursor Activation Before the cofactor is added the precursor protein is called an apoenzyme. Once the cofactor is added it is called a holoenzyme. Sometimes other enzymes can cause the activation eg protease. Other times it is a change in pH or temperature that causes a change in the tertiary structure and activating the percursor enzyme. These types of precursor enzymes are called zymogens or proenzymes. A good example is pepsinogen in the stomach. It is inactive until the change of pH caused by stomach acid transforms it into the active enzyme pepsin which digests proteins.

Poisons and drugs Potassium cyanide inhibits respiration. Protease inhibitors are used to treat infections by viruses. They stop the virus replicating, by preventing it producing a new protein coat. Complete a short essay to explain the action of some poisons and drugs on enzyme activity.

Complete PAG 4.1 The effect of substrate concentration on the rate of an enzyme controlled reaction

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