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BIOL1: Enzymes (8). 3.1.2 The digestive system provides an interface with the environment. Digestion involves enzymic hydrolysis producing smaller molecules.

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Presentation on theme: "BIOL1: Enzymes (8). 3.1.2 The digestive system provides an interface with the environment. Digestion involves enzymic hydrolysis producing smaller molecules."— Presentation transcript:

1 BIOL1: Enzymes (8)

2 3.1.2 The digestive system provides an interface with the environment. Digestion involves enzymic hydrolysis producing smaller molecules that can be absorbed and assimilated.

3  Enzymes as catalysts lowering activation energy through the formation of enzyme substrate complexes.  The lock and key and induced fit models of enzyme action.

4  The properties of enzymes relating to their tertiary structure.  Description and explanation of the effects of temperature, competitive and non- competitive inhibitors, pH and substrate concentration.

5  To take (something) in through.

6  Physiology The conversion of nutrients into living tissue; constructive metabolism.

7  Noun: A substance that increases the rate of a chemical reaction without itself undergoing any permanent chemical change. Or  A person or thing that precipitates an event.

8  The material or substance on which an enzyme acts.

9  To bring about or stimulate the occurrence of; cause.

10  use the lock and key model to explain the properties of enzymes. They should also recognise its limitations and be able to explain why the induced fit model provides a better explanation of specific enzyme properties.

11  Enzymes are globular protein molecules, generally soluble in water.  They have a 3D shape or tertiary structure.  It has hydrophobic amino acid R-groups in the centre of the “ball” and hydrophilic amino acid R-groups around the outside of the ball.

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13  The formation and breakage of glycosidic bonds, ester bonds and peptide bonds need at least 1 enzyme for each.  Process of protein synthesis, digestion, respiration and photosynthesis require a number of different enzymes.  It has been estimated that one cell may contain over 1000 different enzymes.  Most enzymes are made up of 100s of amino acids.

14 1. What do ester bonds hold together? 2. What do glycosidic bonds hold together? 3. What do peptide bonds hold together?

15 1. lipids/Fats. (3 fatty acids + glycerol = triglyceride) 2. Monosaccharides (2 monosaccharides bonded together = disaccharide.) 3. Amino acids. (Bonded together to form a polypeptide)

16  Enzymes that act inside cells = intracellular enzymes.  E.g. - Hydrolases. These are found in lysosomes and hydrolyse (break down) substances cell has taken in by phagocytosis.  - ATPases found inside mitochondria. Involved in synthesis of ATP in aerobic respiration.  Enzymes that act outside cells = extracellular enzymes.  E.g. Digestive enzymes in the alimentary canal. E.g. amylase which hydrolyses starch to maltose.  Enzymes have a precise 3D shape (= tertiary structure).  Have hydrophilic R groups (side chains), on outside of molecules, makes em soluble in water.  Often name of enzyme ends in ase (e.g. ATPase or protease).

17 4. Explain why enzymes are so specific in the reactions that they catalyse. 5. Suggest why all enzymes are protein molecules. 6. The name given to enzymes that catalyse the breakage of peptide bonds is proteases. What name would be given to the enzymes that catalyse the breakage of: (a) Glycosidic bonds (b) Ester bonds

18 4. The catalytic part of the enzyme is the active site. This has a very specific shape. So only one or a few molecules can fit into it. 5. Only proteins can produce such a wide variety of shapes. This is because they are made up from 20 different amino acid sub units. 6. (a) carbohydrases (b) Lipases

19  Heterotrophs obtain their nutrients, (not food), by consuming other organisms. They need to break down the body of the organism they are consuming, digest it, and then absorb it.  Digestion = breaking larger units into smaller, monomer units. So need to break the bonds. Like peptide bonds, ester bonds, and glycosidic bonds. These are catalysed by different types of enzymes.

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21  Some organisms secrete or release enzymes outside their body, onto the food to digest them, then absorb the smaller monomer units.  Others have an internal digestive system. Food taken into body, enzymes mix with it, digests it into monomer units and then it is absorbed. Many of these enzymes are also extracellular (= action takes place outside cells) as they are released from the cells that produce them, onto the food in the digestive system spaces, (like the lumen of your gut).  Also get intracellular = enzymes found in the cytoplasm of the cells or attached to the membranes. Here their action takes place inside cells.

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23 3. Enzymes and protection Many organisms use enzymes as a defence mechanism. E.g. white blood cells called phagocytes take in and digest bacteria using lysosomal enzymes

24 7. Suggest the advantages of having an internal digestive system compared with secreting enzymes outside the organism. 8. Explain why many white blood cells involved in phagocytosis contain a high concentration of enzymes. 9. Suggest why all organisms, no matter what sort of environment they live in, have enzymes in their cells.

25 7. Enzymes are not lost and can be recycled internal system can be regulated to provide the optimum conditions for the enzyme. 8. Their job is to take in and destroy foreign organisms and debris. The destruction is caused by digestive enzymes in the lysosomes of these cells. 9. Enzymes regulate metabolic processes by catalysing reactions at a rate appropriate to the organism.

26 Enzymes are globular proteins, with a specific 3D shape, resulting from the sequence of their amino acids. They are large but only a small region is functional = active site. Usually a cleft or depression, (dip), where another molecule (or substrate) can bind. Substrate is sometimes called the reactants, (the starting materials). Random movements bring the substrate into contact with enzyme.

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28  Substrate fits perfectly into active site and is temporarily bound to some of the R groups of the enzyme’s amino acids. The substrate molecules move around randomly and collide with the active site of the enzyme.

29  When the molecules combine enter a transition state – here bonds in molecules become strained. The molecules are activated. In this state they are more likely to have bonds broken or new ones formed.

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31  One enzyme – one substrate. As only one substrate fits into that enzyme, like a lock and key = the lock-and-key hypothesis. Remember the substrate does not have the same shape as the active site. It has a complementary shape to the active site.

32  In most enzymes when substrate fits into active site, the shape of whole enzyme changes shape, so that it fits around the substrate more closely to give the enzyme-substrate complex. The enzyme can hold the substrate in position for reaction to occur = the induced-fit hypothesis. By altering its shape the enzyme puts a stain on the substrate molecule, which distorts a particular bond on the substrate and so lowers it activation energy needed to break this bond.

33 Once the products are released from the enzyme it returns to its original shape, ready to be used again. Whether by simple lock and key or induced fit the enzyme is specific to this substrate.

34 10. Explain why changing the shape of one of the amino acids that make up the active site, could prevent the enzyme from functioning. 11. Why might changing certain amino acids that are not part of the active site, also prevent the enzyme from functioning?

35 10. The changed amino acid may not bind to the substrate, which will not then be positioned correctly, if at all in the active site. 11. The changed amino acid may be one that forms hydrogen bonds with other amino acids. These bonds will then not form and so affect the tertiary structure of the enzyme. The shape will change, including the active site, so substrate will not fit.

36 Enzyme may catalyse a reaction where: - Substrate is split into 2 or more molecules (Catabolic)

37 Or - 2 or more molecules are joined together (Anabolic). (Amino acid + amino acid = dipeptides)

38 Interactions between the R groups of the enzyme and the atoms of the substrate can break or encourage formation of bonds with the substrate molecule. So 2 or more products are formed. When reaction complete, product(s) leave active site. Enzyme unchanged, so ready to be used again. Pretty quick reactions. E.g. enzyme catalase can bind with hydrogen peroxide ( ‘a super oxide’), split them into water and oxygen, and release products at rate of 10 7 molecules per second.

39  Some slower. Enzyme rubisco involved in photosynthesis can only deal with 3 molecules per second.  Rubisco is the most abundant enzyme on the planet – why?

40 Most of the time the substrate won’t convert into its product without extra energy = activation energy (E A ). = the initial input of energy to start the reaction. Once started reaction continues of its own accord. The role of the enzymes is to lower this activation energy.

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42  Can provide the equivalent of extra heat to a reaction as they increase the energy of the reactants.  Mammals do this by having a constant body temp of  This temp wouldn’t provide enough activation energy for many substrates to work (without enzymes) and if it goes above 40 0 then can permanently damage proteins.

43  Enzymes are the answer as they decrease activation energy. Do this by holding substrate so molecules can react more easily. So reactions with enzymes take place at lower temps than would do without them.

44  Catabolism = substances break down and release energy.  Anabolism = simple molecules are built up into more complex molecules and use energy.

45  Reactions that liberate more energy than they use = exergonic.  Reactions that take in more energy than they liberate = endergonic.

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47 1 – Kinetic energy = matter that is moving and performing work has kinetic energy. 2 – Potential energy = matter that is not performing work but has the ability to do so is potential energy.

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49 12. Describe how the lock-and-key hypothesis of enzyme action differs from the induced fit hypothesis. 13. Explain how enzymes reduce the activation energy of a reaction. 14. Explain why the reduction in activation energy provided by enzymes is essential to living organisms.

50 12. Induced fit = enzyme changes shape to hold the substrate in active site. Lock-and-key = no shape change to enzyme. 13. Enzyme holds the substrate so reaction occurs more easily and requires less activation energy, than the reaction occurring without an enzyme. 14. Most reactions don’t take place at a sufficient rate to sustain life without enzymes. They need to be faster.

51 Catalase is found in tissues of most living things and catalyses breakdown of hydrogen peroxide into water and oxygen. Catalase Hydrogen peroxide → Oxygen + Water Can do this practically. As reaction happens can collect oxygen. Reaction happens quickly at first, and then reaction gets slower and slower until it stops.

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53  Rate of reaction depends on how many enzyme molecules there are and the speed they combine with substrate. At first loads and loads of available enzyme molecules for substrate to bind with. So at beginning of reaction often number of enzyme molecules that limits rate.  As substrate is converted into product there is less and less substrate left. Enzymes molecules are left waiting for substrate. Reaction gets slower and slower then stops. This is where the graph flattens out.

54  So reaction is always fastest at the beginning = called initial rate of reaction.  Can measure this by calculating the slope of a tangent to the curve, as close to time 0 as possible.  Or read off the graph the amount of O 2 given off in the first 30 seconds.

55 15. Why is it better to calculate the initial rate of reaction from a curve, rather than simply measuring how much oxygen is given off in the first 30 seconds?

56 15. Risk of inaccuracy in a single measurement at 30 seconds. Shape of curve gives more accurate value as is based on many readings taken over a given period of time rather than just 1.

57 To compare look at the beginning of reaction as once reaction started amount of substrate begins to vary as substrate is converted to product at different rates. Only at beginning that is def that the differences in reaction rate are caused only by difference in enzyme conc. If double number of enzymes double number of active sites, as long as is plenty of substrate.

58 Easy to measure rate if one of the products if it is a gas. Not always easy though. E.g. looking rate at which amylase breaks down the substrate starch to the product maltose. Both are colourless. Could measure rate starch is used up. Take samples at known times and test with iodine. Could use a colorimeter can measure intensity of blue-black colour. Can plot graph of remaining starch against time. Can then calculate initial reaction rate as before.

59  With this practical it is easier if you mix starch, iodine in potassium iodide solution and amylase and take regular readings of the colour of the mixture in one tube in a colorimeter. Not ideal though as the iodine interferes with the reaction and slows it down.

60  16. Sketch the curve you would obtain if the amount of starch remaining was plotted against time.

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62  Graph below shows results of prac where conc of catalase, (enzyme), was kept constant and conc of hydrogen peroxide, (the substrate), was varied. Curve shows oxygen released against time. Initial rate was calculated for first 30 seconds.  Initial rates of reaction were plotted against substrate conc.

63  The graph shows as substrate conc increases so does the initial rate of reaction. This is cos there are more substrate molecules around. Comes a point where every enzyme active site is occupied and enzyme can’t work any faster. The enzyme is working at its maximum possible rate, known as V max.

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65  At a point in the reaction, (shown on the graph where the line levels off, known as a plateau), either the enzyme conc or the substrate conc prevent any further increase in reaction rate. They are limiting the reaction = limiting factors. If the conc of the limiting factor is increased then the reaction rate increases.

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67 17. Sketch the shape that the graph above would have if excess hydrogen peroxide was not available. 18. Suggest why enzymes are usually maintained at low concentration in cells.

68  18. Enzymes catalyse reaction quickly and can be reused.

69 Graph below shows how the rate varies with temperature. At low temps – slower reaction. This is because molecules have less kinetic energy and so move around slower, so collide less often.

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71  Higher temp – faster rate of reaction. This is due to the molecules having a higher natural kinetic energy and so they vibrate more and move around faster, so collide faster rate, so combining at a faster rate. So when they do collide at higher temp, they have more energy and so have sufficient activation energy to react. It means it is easier for bonds to be broken so reaction can occur.

72  Once temp reached a certain point vibrations of enzyme are so great that as well as making them collide quickly, some of the weaker bonds, (like hydrogen and ionic bonds), which give it it’s precise shape break. Tertiary structure held less and less in shape. Rate of reaction decreases. This happens at approx 45 o in humans. Now the substrate still fits into the active site, but not as well.

73  If enough bonds are broken then tertiary structure is unravelled and enzyme stops working. Enzyme loses shape = denatured. (At approx 65 0 ). The tertiary structure is damaged then it is not reversible. It is too damaged to rebuild. However if only some bonds affected and shape changed to slow reaction but tertiary structure is still intact, then it is often reversible.  Primary structure is not affected.

74  In the human body, above 40 0 substrate molecules fit less well into active site of enzyme and so the rate slows down. As temp goes higher substrate no longer fits so no reactions.

75  Temperature that enzyme catalyse a reaction at its maximum rate = optimum temperature. (For humans = Our body temp is It would be dangerous to maintain a body temp of 40 0, cos even a slight rise in temp would begin to denature enzymes. So you can have a temperature rise when you are ill and your enzymes stay intact.).

76  Mammals and birds are endothermic = able to maintain their internal body temp independently of the environment. Is a cost though = they require greater amount of food than similar sized reptiles.  Other organisms have different optimum temps. Some bacteria that live in hot springs have very high optimum temps – useful in various industrial applications. Some plants have very low ones.

77 19. How would you carry out an experiment to determine the effect of temperature on the rate of breakdown of hydrogen peroxide by catalase? 20. Explain why increased kinetic energy increases the rate of reaction in an enzyme controlled reaction. 21. What type of bonds would you expect to find in greater numbers holding the tertiary structure of heat resistant enzymes, compared with more heat sensitive enzymes? 22. Suggest why the normal body temp of mammals is slightly below the optimum temp of most of the enzymes that occur in the organism.

78 19. Have several catalase – hydrogen peroxide reactions going, at different temperatures. All other factors must remain constant, including the volume & concentration of hydrogen peroxide, (substrate) and the catalase, (enzyme) solutions. Measure the volume of oxygen given off over time. 20. Increased kinetic energy = increased collisions between enzymes and substrates and so increased rate of reactions. 21. Disulphide bonds.

79 22. If normal body temp was too near to the optimum temp for the enzymes then if organism got a fever or exercised, so heat is generated, then enzymes may be denatured. A higher body temp as well as denaturing enzymes also would require more energy.

80 pH = a measure of the hydrogen ion concentration. The higher the conc – the lower the pH, (the more acidic it is). Tertiary structure of enzyme held together by hydrogen and ionic bonds. These bonds are due to the attraction between oppositely charged groups on the amino acids that make up the enzyme. Hydrogen ions, (because of their charge), interfere with hydrogen and ionic bonds, so alters tertiary structure and so changes the active site.

81 Minor changes in pH only affect a few hydrogen and ionic bonds and these can reform if the pH returns to its optimum. This will reduce enzyme activity. Has to be an extreme pH to denature enzyme. All enzymes have an optimum pH, (where they work fastest). For most this is around pH 7. Not all though. Protease, (each enzyme in your stomach), works best at pH 2/3. This is because you have acid in your stomach to kill bacteria that you may have ingested.

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83 23. Individual hydrogen bonds are fairly weak. How can such weak bonds be responsible for holding the tertiary structure of enzymes in place? 24. Enzymes produced by microorganisms are responsible for spoiling food. Using this fact and your knowledge of enzymes, suggest a reason why: (a) Food is heated to a high temperature before canning. (b) Some foods, such as onions, are preserved in vinegar.

84 23. There are many hydrogen bonds. One is weak but together they are strong. 24a. High temps denature enzymes in microorganisms, so they cannot spoil the food. b. vinegar is acidic, so has low pH. The low pH denatures the enzymes in the microorganisms and so preserves the food.

85  Inhibitor = a substance that slows down or stops an enzyme controlled reaction.  Other substances can have the same shape as the substrate and so can fit into the active site of the enzyme. This inhibits the enzyme. Some substances fit onto another part of the enzyme and indirectly cause a shape change n the active site of the enzyme. Most inhibitors affect only one enzyme, others affect many.

86  Reversible Vs irreversible  Some inhibitors can be temporary inhibitors = only binds briefly to active site. In this case if there are loads more substrate molecules than inhibitors, then the substrate can still bind and reaction rate hardly affected.  But if concentration of inhibitor rises or substrate falls then more likely inhibitor will collide with active site of enzyme rather than substrate and so reaction rate slows.

87  Inhibitor can remain in active site permanently. = irreversible inhibition. Even if more substrate added the inhibitor can’t be displaced.

88 (1) If the inhibitor has the same shape as the substrate and can fit into the active site of the enzyme = competitive inhibition. They form an enzyme-inhibitor complex. No product formation cos enzyme does not catalyse a reaction. It is normally reversible. Can be reversed by increasing concentration of substrate.

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91   (2) Sometimes a molecule can bind to another part of the enzyme, rather than the active site. This changes the shape of the active site of the enzyme, by distorting the tertiary structure of the enzyme.

92  Substrate can’t bind, however much of it there is. = non-competitive inhibition. It can be reversible inhibition but more normally is irreversible inhibition. Depending on whether the inhibitor bonds briefly or permanently with the enzyme.

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95  Allosteric = refers to when the enzymes’ shape is altered. When an inhibitor binds with the enzyme on a site other than its active site it changes the shape of the active site, so the substrate can’t bind to it = Non-competitive inhibition.

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97 Whether the inhibition is reversible or non- reversible, whilst the inhibitor is bound to the enzyme the enzyme is said to be denatured. E.g. Digitalis is a substance extracted from the plant foxglove. It is a non- competitive inhibitor. It binds with the enzyme ATPase and results in an increase in the contraction of the heart muscle.

98 25. Suggest a method of determining whether the inhibition of an enzyme- controlled reaction is competitive or non- competitive. 26. Competitive inhibitor molecules can be much larger than the substrate molecules they compete with. Suggest how this is possible.

99 25. Carry out the reaction with a range of substrates concentrations. If the rate of reactions increases up to the same as that given without inhibitor present, then the inhibitor is a competitive inhibitor. 26. The competitive inhibitor may be a large molecule with a small part the same shape as the substrate (and the active site of the enzyme).

100  Some poisons inhibit the enzyme but other over activate the enzyme.

101 Inhibitors that seriously disrupt enzyme controlled reactions can act as metabolic poisons. They prevent vital chemical reactions taking place. E.g. 1. Poison found in the deadly death cap mushroom called alpha- amanitin, inhibits enzymes that catalyse the production of RNA from DNA. Cells can’t make proteins and die.

102  E.g. 2. Potassium cyanide inhibits cell respiration. It is a non-competitive inhibitor for the vital respiratory enzyme cytochrome oxidase, found in mitochondria. ATP can’t be made. The organism can only respire anaerobically which leads to lactic acid build up in the blood. Only need mg for an adult to lose consciousness, in 10s. If untreated go into a coma in 45mins and dead after 2hrs.

103 Found in antifreeze. Not poisonous but when broken down in liver by enzyme alcohol dehydrogenase, the breakdown product called oxalic acid is very toxic and can cause death. Remedy = give massive dose of ethanol, (alcohol). Leads to severe (but less likely to be fatal), alcohol intoxication. The ethanol acts as a competitive inhibitor of alcohol dehydrogenase. This reduces rate of production of oxalic acid, allowing ethylene glycol to be excreted harmlessly.

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105  The antibiotic penicillin works as an inhibitor. It permanently occupies the active site of an enzyme which helps synthesis bacterial cell walls. Prevents new bacterial cells being produced.

106  Snake venom is a mixture of toxins and different enzymes. Phosphodiesterases are in most venoms.  They cause a fall in prey’s blood pressure and cause heart failure.

107 27. Many antibiotics are chemicals that fungi produce and release into their environment. What is the advantage to a fungus of producing and releasing antibiotics? 28. Suggest why proteases inhibitors can inhibit viral proteases, but do not affect the human protease enzymes in the cell.

108 27. The fungi release the antibiotics to destroy other organisms, (bacteria), so that the other organisms cannot take up its food supply. (Defence mechanism). 28. Protease enzymes of viruses differ in shape to that of the protease in humans. Protease inhibitors are specific.

109 Many enzymes need the presence of another non- protein substance to work. These substances = coenzymes, or cofactors. No real difference between the 2 but some use cofactor to mean simple molecule inorganic ion and term coenzyme for bigger molecules. Both work by binding briefly with enzyme. Sometimes this alters their shape so can bind more effectively with substrate. Sometimes it helps the enzyme to transfer a particular group of atoms from one group to another.

110  Small, inorganic non-protein molecules. Bind for a short time to the active site, either just before or at same time as substrate. In many reactions the coenzymes take place in the reaction and like substrates are changed during it. Unlike substrates though, they can be recycled back again though.  Many cofactors are made from vitamins. E.g. vitamin B 3 (nicotinamide) is important role in breaking down fats and carbohydrates. Vitamin B 3 used to make a coenzyme required for the enzyme pyruvate dehydrogenase to function properly. This enzyme is required in respiration.  Deficiency in B 3 leads to disease known as pellagra. This causes diarrhoea, dermatitis, dementia and death eventually.

111 - = a coenzyme which is permanent part of an enzyme. They contribute to final 3D shape of enzyme. E.g. the enzyme carbonic anhydrase contains zinc based prosthetic groups. Enzyme essential component of red blood cells. Involved in catalysing the combining of carbon dioxide and water to form carbonic acid. (Which is how CO 2 transported in blood).

112 small inorganic ions. May combine with enzyme or substrate. The binding makes the enzyme-substrate complex form more easily cos it affects the charge distribution and sometimes the shape of the enzyme-substrate complex. E.g. (cofactor) chloride ions bind to salivary amylase, changes its shape a bit and help starch to bind to if more efficiently. Starch is then broken down into maltose. E.g.2 (cofactor) calcium ions essential for enzyme thrombin, which catalyses change of soluble, globular protein fibrinogen into the insoluble, fibrous protein fibrin, used in blood clotting. E.g. 3 (coenzyme) = coenzyme A. needed in many metabolic pathways including aerobic respiration.

113 29 Suggest why the recommended daily dietary allowance for nicotinamide (18mg), is very low. 30. Name the prosthetic group found in haemoglobin. 31. Suggest where in the cell the addition of prosthetic groups to the enzyme molecules take place.

114 31. It is a coenzyme and is reusable. 32. The haem group. 33. The Golgi apparatus.

115 They are specific so they do not produce a range of unwanted products hence they have a broad use in industry:  28% detergents  35% food processing  23% beverages  14% others


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