Presentation MEDIA: Genetics & Evolution Series. The Nature of Genes. Set No. 3

Transcription

1 Presentation MEDIA: Genetics & Evolution Series The Nature of Genes Set No. 3

2 Presentation MEDIA Biozone International Ltd ISBN OHT Title 1 Metabolism Index to OHT Titles 2 The Concept of the Gene 3 Gene Expression 4 Enzymes 5 Enzyme Structure 6 Types of Enzyme 7 Mechanism of Enzyme Action 8 Enzymes are Catalysts 9 ph Effects on Enzyme Activity 10 Temperature Effects on Enzyme Activity 11 Enzyme Concentration Effects on Enzyme Activity 12 Substrate Concentration Effects on Enzyme Activity 13 The Effect of Cofactors on Enzyme Activity 14 The Nature of Enzyme Inhibitors 15 Reversible Enzyme Inhibitors 16 Irreversible Enzyme Inhibitors 17 Anabolism 18 Catabolism 19 Regulation of Metabolism 20 Location of Enzymes and their Substrates OHT Title 21 The Arrangement of Enzymes within Cells 22 Metabolic Pathways 23 Metabolism of Phenylalanine 24 Inherited Metabolic Disorders 1 25 Inherited Metabolic Disorders 2 26 Inherited Metabolic Disorders 3 27 Regulation of Enzyme Production 28 Structure of the Operon 1 29 Structure of the Operon 2 30 Operon Function in Prokaryotes 31 Gene Induction 1 32 Gene Induction 2 33 Gene Induction 3 34 Gene Repression 1 35 Gene Repression 2 36 Gene Repression 3 37 Gene Induction and Repression - Summary 38 Control of Gene Expression in Eukaryotes 1 39 Control of Gene Expression in Eukaryotes 2 40 Control of Metabolism 1 41 Control of Metabolism 2 NEW ZEALAND: Biozone International Ltd P.O. Box Hamilton Telephone: +64 (7) FAX: +64 (7) info@biozone.co.nz AUSTRALIA: Biozone Learning Media Australia P.O. Box 7523 GCMC 4217 QLD Telephone: +61 (7) FAX: +61 (7) info@biozone.com.au UNITED KINGDOM: Biozone Learning Media (UK) P.O. Box 16710, Glasgow G12 9WS Telephone: +44 (141) FAX: +44 (141) info@biozone.co.uk

3 Metabolism The life processes of a cell are a vast collection of chemical reactions called metabolism. Definition: Metabolism The chemical activities of life. All the various processes by which you: obtain energy grow heal think feel dispose of waste materials The average adult human eats 1 tonne of food per year, yet body composition and size usually stays much the same. Therefore a tremendously complex series of chemical reactions must take place to 'maintain' the organism. Failure of a single enzyme can lead to collapse of a system and death! Metabolism OHT 1

4 The Concept of the Gene Metabolism is controlled by a vast assortment of enzymes. Genes code for the production of proteins, many of which are enzymes. The activity of genes can be regulated this controls the production of enzymes. Other factors can regulate the activity of enzymes after production. Gene expression (activity) is regulated by other 'controller genes' Enzyme activity is controlled by a number of factors Transcription Translation Protein DNA Contains 'blueprint' for the manufacture of all proteins mrna Some proteins are enzymes that control the metabolic processes of the cell OHT 2

5 OHT 3 DNA DNA contains the master copy of all the genetic information to produce proteins for the cell Reverse transcription occurs when retroviruses invade host cells. Their viral RNA is converted to DNA and spliced into the host's genome Transcription Gene Expression Genes code for the production of proteins, many of which are enzymes: mrna mrna is an exact copy of part of the DNA molecule coding for making a single protein Amino Acid Translation Tyr trna is a carrier molecule responsible for bringing in the amino acids in their correct sequence to make a protein trna Protein Structural? Regulatory? Contractile? Immunological? Transport? Catalytic?

6 Enzymes Most enzymes are proteins. Enzymes act as biological catalysts (regulating cell metabolism). The molecule that an enzyme acts on is called the substrate. Enzymes are specific for the reactions they catalyse. Enzyme activity depends on the shape of the enzyme and the arrangement of its active site (the binding site for the substrate). Many enzyme names end in -ase (but not all). EXAMPLES: Lipase Trypsin Amylase Renin Lactase Cholinesterase Breaks down fats Breaks down proteins Breaks down starch Coagulates milk protein Breaks down milk sugar Breaks down the neurotransmitter acetylcholine in the nervous system OHT 4

7 Enzyme Structure Substrate molecule is the chemical (in this case RNA) that an enzyme acts on RNA Active sites are attraction points that draw the substrate to the surface of the enzyme Enzyme Molecule is a specific catalyst. The complexity of the active site is what makes each enzyme so specific (i.e. precise in terms of the substrate it acts on) Source: Lubert Stryer Ribonuclease S (shown above) is an enzyme that breaks up RNA molecules. The dark (red) areas form the active site and are comprised of certain amino acid 'R' groups. The substrate (RNA) is drawn into the active site, putting the substrate molecule under stress, thereby causing the reaction to proceed more readily. OHT 5

8 Types of Enzyme Nearly all enzymes are made of protein, although RNA has been shown to have enzyme properties. Some enzymes consist of just protein, while others, called conjugated protein enzymes, require additional components to complete their catalytic properties. These may be permanently attached parts called prosthetic groups, or temporarily attached pieces (coenzymes) that detach after a reaction, and may participate with another enzyme in other reactions. Protein-only Enzymes Active site enzyme Enzyme comprising just protein e.g. Lysozyme Conjugated Protein Enzymes Apoenzyme Active site Prosthetic group is more or less permanently attached Active site Apoenzyme Coenzyme becomes detached after the reaction Prosthetic Group Required Contains apoenzyme (protein) plus a prosthetic group e.g. Flavoprotein + FAD Coenzyme Required Contains apoenzyme (protein) plus a coenzyme (non-protein) e.g. Dehydrogenases + NAD OHT 6

9 Mechanism of Enzyme Action Steps in an Enzyme's Activity An enzyme and its substrate act somewhat like a lock and key. The shape of the enzyme changes when the substrate fits into the cleft (called the induced fit): 1 Two substrate molecules are drawn into the cleft of the enzyme Enzyme Cleft Substrate molecules 2 The enzyme changes shape, forcing the 2 substrate molecules to combine Enzyme Enzyme changes shape 3 The resulting end product is released by the enzyme, which returns to its normal shape, ready to receive more Enzyme End product released The specificity of the substrate is determined by the complexity of the binding sites (this can be absolute). Some enzymes have specificity to a bond type (e.g. lipases break up any chain length of lipid). OHT 7

10 Amount of Energy Stored in the Chemicals Enzymes are Catalysts Enzymes act as biological catalysts. They alter the chemical equilibrium between reactant and product. When the substrate attains the required energy it is able to change into the product or products. All catalysts speed up reactions by: Influencing the stability of bonds in the reactants. Providing an alternative reaction pathway: the binding of reactants and enzyme can weaken bonds in the reactants and allow the reaction to proceed more easily. High Reactant High Energy Without Enzyme: The activation energy needed to make the reaction proceed in the forward direction is high without the enzyme present With Enzyme: The energy required for the reaction to proceed is reduced by the presence of the enzyme. Reactants turn into products more readily. Product Low Start Direction of Reaction Low Energy Finish OHT 8

11 ph Effects on Enzyme Activity Like all proteins, enzymes are denatured (made nonfunctional) by extremes of ph (acidity/alkalinity). Within these extremes most enzymes are still influenced by ph. There is a particular ph for optimum activity for each enzyme. This is because the active sites of the enzyme can be destroyed by the wrong ph. Optimum ph for urease Optimum ph for trypsin Enzyme Activity Pepsin Urease Trypsin Acid ph Alkaline OHT 9

12 Temperature Effects on Enzyme Activity Speeds up all reactions, but the rate of denaturation of enzymes also increases at higher temperatures. High temperatures break the disulphide bonds holding the tertiary structure of the enzyme together. This destroys the active sites and therefore makes the enzyme non-functional. Optimum temperature for enzyme Enzyme Activity Too cold for the enzyme to operate Rapid denaturation Temperature ( C) OHT 10

13 Enzyme Concentration Effect on Enzyme Activity Assuming that the amount of substrate is not limiting, an increase in enzyme concentration causes an increase in the reaction rate. Rate of Reaction With ample substrate and cofactors present Enzyme Concentration OHT 11

14 Substrate Concentration Effect on Enzyme Activity Assuming that the amount of enzyme is constant, an increase in substrate concentration causes a diminishing increase in the reaction rate. A maximum rate is obtained at a certain concentration of substrate when all enzymes are occupied by substrate (the rate cannot increase any further). Rate of Reaction With ample enzyme and cofactors present Concentration of Substrate OHT 12

15 Effect of Cofactors on Enzyme Activity Cofactors are substances that are essential to the catalytic activity of some enzymes. Cofactors may alter the shape of enzymes slightly to make the active sites functional or to complete the reactive site. Enzyme cofactors include coenzymes (organic molecules) and activating ions (e.g. Na +, K +, Ca 2+, Mg 2+ ). Vitamins are often coenzymes. Vitamins are organic molecules not synthesised by the body. e.g. Vitamin K, vitamin B1, vitamin B6, folic acid) Enzyme Once the shape of the enzyme has been modified by the cofactor, substrates A and B can react together A B The presence of the cofactor alters the shape of the enzyme Enzyme A B Product OHT 13

16 The Nature of Enzyme Inhibitors Enzyme inhibitors may or may not act reversibly: Reversible: The inhibitor is temporarily bound to the enzyme, thereby preventing its function (used as a mechanism to control enzyme activity). Irreversible: The inhibitor may bind permanently to the enzyme causing it to be permanently deactivated. Enzyme inhibitors work in one of two ways: Competitive inhibitors: The inhibitor competes with the substrate for the active site, thereby blocking it and preventing attachment of the substrate. Noncompetitive inhibitors: The inhibitor binds to the enzyme (but not at the active site) and alters its shape. It markedly slows down the reaction rate by making the enzyme less able to perform its function. Allosteric enzyme inhibitors are non competitive inhibitors that block the active site altogether, preventing the substrate from binding. In this case, the enzyme ceases to function. OHT 14

17 Reversible Enzyme Inhibitors Enzyme inhibitors may be of two kinds: reversible and irreversible. Reversible inhibitors are used to control the activity of an enzyme. There is often an interaction between the substrate or end product and the enzyme controlling the reaction. Buildup of the end product or a lack of substrate may serve to deactivate the enzyme. This deactivation may take the form of competitive (competes for the active site) or noncompetitive inhibition. Substrate No inhibition Good fit Enzyme Competitive inhibitor blocks the active site Enzyme Substrate The inhibitor binds to the enzyme, and alters the enzyme s ability to function properly The substrate binds to the active site Enzyme Competitive inhibitor Noncompetitive inhibitor OHT 15

18 Irreversible Enzyme Inhibitors Irreversible enzyme inhibitors are poisons that prevent an enzyme from catalysing a reaction. Heavy metals: Certain heavy metals bind tightly and permanently to the active sites of enzymes, destroying their catalytic properties. Examples of toxic heavy metals include: mercury (Hg), cadmium (Cd), lead (Pb), and arsenic (As). They are generally non-competitive inhibitors, although an exception is mercury which deactivates the enzyme papain. Heavy metals are retained in the body, and lost slowly. Substrate The substrate cannot lock on to the active sites The inhibitor blocks the active sites Hg Enzyme Active sites Insecticides These can prevent the breakdown of acetylcholine (ACh), a neurotransmitter in the nervous system. They bind to the enzyme that normally breaks down the ACh, causing over stimulation of the nerves. OHT 16

19 OHT 17 Substrate A Enzyme 1 Active Sites Substrate molecules enter the enzyme active site Substrate B Anabolism Anabolism is the build up or synthesis of complex molecules from simpler ones to make chemicals needed by the cell. This process requires energy. Examples: 1. Protein synthesis: proteins are made from amino acids. 2. Photosynthesis: sugar (glucose) is made from water and carbon dioxide with the input of light energy. 2 Substrate subjected to stress which aids the formation of bonds 3 Product Substrate molecules form a single product which is released

20 OHT 18 Enzyme Substrate 1 Substrate enters the active sites Active Sites Catabolism Catabolism is the break down of complex, high energy, molecules into simpler ones with lower energy. This process releases energy, including heat to keep us warm. Examples: 1. Digestion of food: carbohydrates, proteins, and fats are broken down into their building blocks for absorption. 2. Cellular respiration: glucose molecules are broken down to release energy. 2 Substrate is subjected to stress facilitating the breaking of bonds Product A 3 Product B Substrate is broken in two and the products are released

21 Regulation of Metabolism The overall activity of enzymes, and therefore metabolism, is controlled by a number of factors: The rate of enzyme production (by protein synthesis) The rate of enzyme breakdown (degradation) The influence of cofactors and inhibitors Changing the activity of the enzyme by its interaction with the substrate or the products of the reaction it is controlling: Speed forward stimulation (interaction with substrate) Negative feedback (interaction with product) Speed Forward Stimulation operates where the substrate must be kept at a low concentration Negative Feedback operates where large amounts of the end product deactivates enzyme 1 at the beginning of the metabolic pathway Enzyme 1 Enzyme 2 Substance A Substance B Substance C Substrate (starting chemical) End product (finishing chemical) OHT 19

22 Location of Enzymes and their Substrates Enzymes are often located in specific regions of the cell e.g. in mitochondria or chloroplasts. This results in greater cell efficiency since the enzymes for a particular metabolic pathway (e.g. the respiratory chain enzymes in the mitochondria) can all be kept within a single type of organelle. The rate of enzyme reaction in these cases is partly determined by the rate at which substrates can enter the organelle through the cell membrane. Outer membrane Inner membrane Mitochondrial DNA Ribosome Matrix Cristae OHT 20

23 The Arrangement of Enzymes within Cells Enzymes do not always exist in isolation. They are often grouped together and bound to the inner surface of membranes e.g. in the mitochondria. The enzymes are assembled together to catalyse several steps of a metabolic pathway. The spatial arrangement of the enzymes orders the sequence of reactions, since the product of one reaction is the substrate for the next. Amine oxidases and other enzymes on the outer membrane surface Matrix Adenylate kinase and other phosphorylases between the membranes Respiratory assembly enzymes embedded in the membrane (ATPase) Cross-section of a mitochondrion Many soluble enzymes of the citric acid cycle floating in the matrix, as well as enzymes for fatty acid degradation OHT 21

24 OHT 22 Precursor Chemical Protein synthesis produces enzyme A Metabolic Pathways A metabolic pathway is a series of steps from a starter molecule or precursor toward a final end product. Each step is catalysed by a different enzyme whose structure is coded by a specific gene (one gene codes for one enzyme). Gene A Enzyme A Enzyme A transforms the precursor chemical into the intermediate chemical by altering its chemical structure Intermediate Chemical Gene B Enzyme B Protein synthesis produces enzyme B Enzyme B transforms the intermediate chemical into the end product End Product

25 Metabolism of Phenylalanine The scheme below shows the conversion of the essential amino acid phenylalanine into many derived products. The failure of specific enzymes causes metabolic disorders: Protein Phenylketonuria Proteins are broken down to release free amino acids one of which is phenylalanine Thyroxine a series of enzymes Phenylalanine Phenylalanine hydroxylase Tyrosine Faulty enzyme causes buildup of: Tyrosinase This in turn causes: Phenylpyruvic acid Melanin Cretinism Faulty enzymes cause: Transaminase Faulty enzyme causes: Albinism Hydroxyphenylpyruvic acid Hydroxyphenylpyruvic acid oxidase Faulty enzyme causes: Tyrosinosis Homogentisic acid Homogentisic acid oxidase Faulty enzyme causes: Alkaptonuria Maleylacetoacetic acid Carbon Dioxide and Water OHT 23

26 Inherited Metabolic Disorders 1 The symptoms of the various disorders associated with the faulty metabolism of phenylalanine are: Phenylketonuria: Mental retardation, 'mousy body odour', light skin colour, excessive muscular tension and activity, eczema. Cretinism: Dwarfism, mental retardation, low levels of thyroid hormones, retarded sexual development, yellow skin colour. Albinism: Complete lack of the pigment melanin in body tissues, including skin, hair and eyes. Tyrosinosis: Death from liver failure, or (if surviving) chronic liver and kidney disease. Alkaptonuria: Dark urine, pigmentation of cartilage and other connective tissues, and in later years - arthritis. OHT 24

27 Inherited Metabolic Disorders 2 Most inherited metabolic disorders are caused by faulty enzymes, such as the ones listed below: Phenylketonuria (PKU) Caused by: Faulty gene results in the absence of an enzyme in the liver, allowing phenylalanine to rise to harmful levels. Leads to: Brain damage. Occurrence: 1 in 19,400 newborn babies Cystic Fibrosis Caused by: Abnormal control of secretions (body fluids). Leads to: Poor growth, chest infections, shortened life. Occurrence: 1 in 4,100 newborn babies Maple Syrup Urine Disease (MSUD) Caused by: Non-functional enzyme (3 amino acids involved). Leads to: Life-threatening complications. Occurrence: 1 in 166,500 newborn babies Galactosemia Caused by: Enzyme defect prevents normal use of milk sugar. Leads to: Jaundice, cataracts, and severe illness. Occurrence: 1 in 67,600 newborn babies OHT 25

28 Inherited Metabolic Disorders 3 Additional examples of inherited metabolic disorders caused by faulty enzymes: Congenital Hypothyroidism Caused by: Leads to: Not enough normal thyroid gland. Slowed growth and mental development. Occurrence: 1 in 4,900 newborn babies Congenital Adrenal Hyperplasia Caused by: Leads to: A deficiency in a group of hormones which regulate salt balance and levels of testosterone (male sex hormone) in either sex. Vomiting, dehydration and death within days. In girls: genitalia appear masculine, despite female internal sex organs. In boys: early development of secondary sex characteristics occurs. Occurrence: 1 in 22,900 newborn babies Biotinidase Deficiency Caused by: Leads to: A deficiency of 3 mitochondrial enzymes. Neurological disorder and low ph of body fluids possibly leading to coma. Occurrence: 1 in 22,900 newborn babies OHT 26

29 Regulation of Enzyme Production Cells need to control the rate and frequency of protein synthesis. These controls often occur at transcription. Sometimes genes are induced (transcribed) only when an enzyme product is required to catalyse reactions that may occur infrequently e.g. use of a particular substrate that is not always available. Other genes are being transcribed all the time because their enzyme products are in constant demand e.g. the genes coding for respiratory enzymes. In prokaryotes operons control the rate of transcription. An operon is a group of closely related genes that act together and code for the enzymes regulating a particular metabolic pathway. Transcription Translation DNA Transcription stage may be switched ON or OFF mrna Enzyme OHT 27

30 Structure of the Operon 1 The operon in prokaryotes comprises a number of different genes: 1. Structural Genes These code for the production of the enzymes involved in a particular set of reactions. e.g. - Use of lactose as a carbohydrate source. - Production of the amino acid tryptophan. 2. Promoter Gene This is the recognition site for the RNA polymerase enzyme to bind to. 3. Operator Gene Controls the production of mrna. Located outside the operon DNA strand Regulator Gene Promoter Operator Structural Gene A OPERON The operon consists of the structural genes Outside the operon, a regulator gene produces a repressor molecule which can block the operator gene. OHT 28

31 OHT 29 Protein Synthesis Regulator Gene The regulator gene, on another part of the DNA, produces the repressor molecule by protein synthesis Structure of the Operon 2 RNA polymerase The RNA polymerase enzyme creates a mrna copy of the structural genes to intitiate protein synthesis Progress may be blocked Promoter The promoter site is where the RNA polymerase enzyme first attaches itself to the DNA to begin synthesis of the mrna An active repressor molecule will bind to the operator site Repressor Operator Structural Gene A The operator is the potential blocking site. It is here that an active repressor molecule will bind, stopping mrna synthesis from proceeding. OPERON At least one structural gene is present which codes for the creation of an enzyme in a metabolic pathway DNA The operon consists of the structural genes and the promoter and operator sites

32 OHT 30 Regulator Gene Operon Function in Prokayotes Two alternative processes are thought to control the activity of operons: 1. Repression: where gene transcription is switched OFF 2. Induction: where gene transcription is switched ON A series of enzyme controlled reactions transforms the substrate into the end product. At each step, the chemical is altered slightly in its chemical makeup Protein Synthesis RNA polymerase Progress may be blocked Promoter Substrate An active repressor molecule will bind to the operator site Repressor Operator Structural Gene A Metabolic Pathway Structural Gene B Translation Transcription Structural Gene C Structural Gene D End Product Chemical 1 Chemical 2 Chemical 3 Chemical 4 Chemical 5 Enzyme A Enzyme B Enzyme C Enzyme D OPERON Functional enzymes mrna DNA

33 Gene Induction 1 In this type of gene regulation the genes are normally switched off but are switched on when they are required. A well studied operon of this type in bacteria is the lac operon in E. coli (the prefix lac refers to the substrate involved which is lactose). Step 1: The Production of the Repressor Protein The regulator gene is on a distant part of the chromosome from the operon and produces a protein, called a repressor molecule. In the absence of the substrate (lactose), the repressor can block the binding site of the synthesiser enzyme (RNA polymerase). This prevents transcription of the genes coding for enzyme synthesis (when lactose is not available). The regulator gene on another part of the chromosome produces a protein called a repressor molecule Repressor moves to block operator RNA polymerase Repressor Transcription is prevented by the repressor molecule blocking the operator site so that the RNA polymerase cannot transcribe the structural genes. Regulator Gene Promoter Operator Structural Gene A OHT 31

34 Gene Induction 2 When the substrate (lactose) is present, and therefore needs metabolising, it may act as an inducer molecule. Step 2: The Inducer Binds to the Repressor Protein This is a reversible reaction that will only happen if the inducer molecule (the substrate, lactose), is in high concentration. The inducer binds to the repressor protein preventing it from binding to the RNA polymerase binding site. RNA polymerase can then bind and the structural genes can be transcribed. The inducer molecule may be the substrate (precursor molecule) for the beginning of the metabolic pathway Inducer The repressor protein may be approached by an inducer molecule and bind to it (this is a reversible reaction that will only happen when the inducer molecule is in high concentration) Inducer binds to the repressor altering its shape so it is no longer able to bind to the DNA Inducer Repressor Repressor Regulator Gene Promoter Operator Structural Gene A OHT 32

35 Gene Induction 3 Step 3: Gene Transcription and Enzyme Synthesis Once the repressor protein is deactivated the synthesiser enzyme (RNA polymerase) can get access to the operator gene. mrna is transcribed in one continuous piece, coding for all of the structural genes in the operon. The enzymes are produced in a sequence that reflects the stages in the metabolic pathway that they code for. As one enzyme is created by protein synthesis then the RNA polymerase moves to the next structural gene. This type of regulatory system allows the cell to make enzymes only when there is sufficient substrate (i.e. their production is induced by the presence of the substrate). RNA polymerase produces one continuous piece of mrna for all the structural genes in the operon Promoter Operator Structural Gene A Structural Gene B With the repressor removed, RNA polymerase can get access to the operator gene RNA polymerase OHT 33

36 Gene Repression 1 Another type of gene regulation occurs where the operon is normally switched on. The genes are turned off only when the end product is present in large quantities. Example: tryptophan operon in E.coli. An effector molecule is required to activate the repressor. The effector molecule is usually the end product of a metabolic pathway. Step 1: The Repressor is at First Inactive When the effector (end product) is in low concentration the repressor molecule is the wrong shape and cannot bind to the operator site. Transcription of the structural genes is not blocked. The regulator gene on another part of the chromosome produces a protein called a repressor molecule The repressor molecule in this form is inactive and is incapable of binding to the operator site to block mrna synthesis Repressor Transcription using the RNA polymerase enzyme proceeds uninterrupted Regulator Gene Promoter Operator Structural Gene A RNA polymerase OHT 34

37 Gene Repression 2 Step 2: The Repressor is Activated When the effector (end product) is in high concentration it binds to the repressor and changes its shape. Effector in high concentration Effector Repressor The repressor molecule has its shape changed as the effector molecule binds to it. This only occurs when the effector is in high concentration Effector Repressor OHT 35

38 Gene Repression 3 Step 3: The Repressor Binds to the Operator The shape change of the repressor molecule enables it to bind to the operator. As a result, transcription is switched off. The structural genes cannot be transcribed because the RNA polymerase cannot bind to the promoter site. RNA polymerase RNA polymerase is prevented from binding to the promoter site to begin transcription Effector The now active repressor molecule is able to bind to the operator site and prevent transcription Promoter Repressor Operator Structural Gene A OHT 36

39 Gene Induction and Repression Summary In prokaryotes, genes can occur as operons which can be switched on or off by regulating genes. There are two types of operon recognised: Gene Induction Genes that are induced are normally switched off. The inducer is the substrate that becomes available e.g. as an energy source. The presence of the substrate deactivates the repressor allowing transcription of structural genes to proceed. Gene Repression Genes that are repressible are normally switched on. The presence of high levels of the end product of a metabolic process activates the repressor molecule. The active repressor prevents further transcription of the structural genes. OHT 37

40 Control of Gene Expression in Eukaryotes 1 The control of gene expression in eukaryotes is similar in nature, but more complex than that in prokaryotes: Activators are proteins that bind to enhancer genes and help: to determine which genes will be switched on speed up the rate of transcription Repressors bind to selected sets of genes known as silencers: they interfere with the functioning of activators they slow down the rate of transcription Enhancer Silencer Repressor DNA Molecule Enhancer Activator Activator Activator Enhancer Basal Factors position RNA polymerase enzyme at the start of the gene TATA Binding Protein RNA Polymerase mrna start site TATA Coactivators are adapter molecules that receive signals from the activators and repressors and pass them on to the basal factors Source: Scientific American, Molecular Machines That Control Genes, February 1995, pp TATA box is located about 30 base pairs from the mrna start site Promoter Region A regulator region a short distance from the beginning of a gene that acts as a binding site for the RNA polymerase enzyme Protein Coding Region of a gene where it begins to transcribe the DNA code into mrna OHT 38

41 Control of Gene Expression in Eukaryotes 2 The control of gene expression in human cells is carried out by regulating the process of transcription. Basal factors are essential for transcription but cannot by themselves increase or decrease its rate. Transcription rate is regulated by activators and repressors and these can vary from gene to gene. Activators attach to parts of the DNA called enhancer regions and act to speed up the rate of transcription. In contrast, repressors attach to silencer regions of the DNA and act to slow down transcription. Activators communicate with the basal factors through coactivators. These proteins are linked to the TATA binding protein. The first of the basal factors to land on the region of genes known as the core promoter. When the basal factors are stimulated by the presence of activators, RNA polymerase is positioned at the start of the coding sequence and begins transcription. Because each gene has a unique combination of silencers and enhancers, cells can control transcription of every gene individually. OHT 39

42 Control of Metabolism 1 Organisms have many control mechanisms involved in regulating metabolism. These can occur at: 1. The DNA level 2. At the transcriptional level 3. When mrna is being translated 4. After the protein is made Control at the DNA Level: 1. Gene deactivation: In eukaryotes, chromatin sometimes remains packed up and is not transcribed e.g. Barr bodies in humans. 2. There may be multiple copies of genes coding for products required in high levels. Transcriptional Control: 1. Original mrna molecules can be modified by the removal of meaningless sequences. 2. Genes can be switched on or off with repressors. Post-Transcriptional Control: 1. The rate of ribosome attachment and detachment controls the speed of translation. 2. The rate of translation can be controlled by the length of life of the mrna molecule. OHT 40

43 Control of Metabolism 2 Post-Translational Control: 1. The rate of enzyme breakdown (degradation) controls the amount of an end product. 2. Feedback inhibition can prevent the functioning of enzymes in the initial steps of a metabolic pathway. 3. Enzymes may be synthesised in an inactive form e.g. protein digesting enzymes that would be dangerous if stored in the active form. 4. Proteins can be modified by the addition of other molecules e.g. carbohydrates, which alter the function of the protein. Cell Compartments 1. Enzymes can be restricted within cells to different organelles e.g. respiratory enzymes in mitochondria. The reaction rate will be limited partly by the speed at which substrates enter the organelle. 2. Specific types of enzymes are often found in certain organs e.g. enzymes catalysing the reactions of the urea cycle are found mainly in the liver. OHT 41