Week 9- Enzymes PDF

ENZYMES LET US SPEED IT UP WHAT ARE ENZYMES An enzyme is a biocatalyst, which enhances the rate of thermodynamically favorable biological reactions to several thousand to million folds Importance of Enzymes Nothing works without enzymes !!!  CATALYST chemical...

ENZYMES LET US SPEED IT UP WHAT ARE ENZYMES An enzyme is a biocatalyst, which enhances the rate of thermodynamically favorable biological reactions to several thousand to million folds Importance of Enzymes Nothing works without enzymes !!!  CATALYST chemical that speeds up the reaction but is not used up in the reaction ACTIVATION ENERGY the required input of energy to make a reaction start  Properties of Enzymes Most enzymes are three dimensional globular proteins (tertiary and quaternary structure)  Some special RNA species enzymes and are called Ribozymes  General Properties of Enzymes 1. They accelerate the reaction, but a. do not alter the reaction equilibrium b. not consumed in overall reaction c. required only in very small quantities. 2. They have enormous power for catalysis. 3. Enzymes are highly specific for their substrate. 4. Enzymes possess active sites  Active Site The active site of an enzyme is a specialized region where a substrate can fit precisely. This microenvironment facilitates and accelerates chemical reactions by providing optimal conditions for substrate conversion. The amino acids R-groups (side  Substrate molecule that an enzyme reacts with and is the substance that binds to the active site of an enzyme. Enzymes are specific to their substrates The specificity is determined by the active site Enzyme Structure  Enzyme Definition Apoenzyme Coenzyme (Cofactor) Holoenzyme–The Protein part of a Non-protein part of a biochemically active conjugated enzyme conjugated enzyme conjugated enzyme. Enzyme Definition  Enzyme Nomenclature Enzymes are named according to the type of reaction they catalyze and/or their substrate Some digestive Suffix of an enzyme– enzymes have the ase suffix –in -Lactase, amylase, –Pepsin, trypsin& Prefix denotes the type of lipase or protease chymotrypsin reaction the enzyme catalyzes –Oxidase: redox reaction –Hydrolase: Addition of water to break one component into two parts 6 Major Classes of Enzymes  EC 1. Oxidoreductases Biochemical Activity: – Catalyse Oxidation/Reduction Reactions Act on many chemical groupings to add or remove hydrogen atoms. Examples: – Oxidase - oxidation of substrate – Reductase - reduction of substrate – Dehydrogenase-introduction of double bond (oxidation) by formal removal of two hydrogen atoms from substrate, the H being accepted by a coenzyme.  EC 2. Transferases Biochemical Activity: Transfer a functional groups (e.g. methyl or phosphate) between donor and acceptor molecules. Examples:  Transaminases - transfer of an amino group between substrates.  Kinases -transfer of a phosphate group between substrates.  EC 3. Hydrolases Biochemical Activity: – Catalyse the hydrolysis of various bonds Add water across a bond. Examples: – Lipase - hydrolysis of ester linkages in lipids – Proteases- hydrolysis of amide linkages in proteins. – Nucleases - hydrolysis of sugar phosphate ester bonds in nucleic acids. – Carbohydrases - hydrolysis of glycosidic bonds in carbohydrates. – Phosphatases - hydrolysis of phosphate- ester bonds.  EC 4. Lyases Biochemical Activity:  Cleave various bonds by means other than hydrolysis and oxidation.  Add Water, Ammonia or Carbon dioxide across double bonds, or remove these elements to produce double bonds. Examples:  Dehydratases - removal of H20 from substrate  Decarboxylases -removal of CO2 from substrate o Deaminases - removal of NH3 from substrate  Hydratase - addition of H2O to a substrate  EC 5. Isomerases Biochemical Activity: – Catalyse isomerization changes within a single molecule. Examples:Mutase reactions (Shifts of chemical groups). – Racemases - conversion of D to L isomer or vice versa. – Mutases - transfer of a functional group from one position to another in the same molecule. – Isomerase -catalyze reactions involving a structural rearrangement of a molecule.  EC 6. Ligases Biochemical Activity: – Join two molecules with covalent bonds Catalyse reactions in which two chemical groups are joined (or ligated) with the use of energy from ATP. Examples: – Synthetases formation of new bond between two substrates with participation of ATP. – o Carboxylases - formation of new bond between a substrate and CO2 with participation of ATP.  Enzyme –Substrate Complex When the substrate binds to the enzyme active site an Enzyme-Substrate Complex is formed temporarily –Allows the substrate to undergo its chemical reaction much faster Lock & Key Model of Enzyme Action The active site is fixed, with a rigid shape (LOCK) The substrate (KEY) must fit exactly into the rigid enzyme (LOCK) Key (substrate) fits into the lock (enzyme) Upon completion of the chemical reaction, the products are released from the active site, so the next substrate molecule can bind Induced Fit Model of Enzyme Action Many enzymes are flexible & constantly change their shape The shape of the active site changes to accept & accommodate the substrate Conformation change in the enzyme’s active site to allow the substrate to bind SPECIFICITY OF ENZYMES  Absolute Specificity An enzyme will catalyze a particular reaction for only one substrate Most restrictive of all specificities (Not common) –Catalase has absolute specificity for hydrogen peroxide (H2O2) –Urease catalyzes only the hydrolysis of urea  Group Specificity The enzyme will act only on similar substrates that have a specific functional group Carboxypeptidase cleaves amino acids one at a time from the carboxyl end of the peptide chain Hexokinase adds a phosphate group to hexoses  Linkage Specificity –The enzyme will act on a particular type of chemical bond, irrespective of the rest of the molecular structure –The most general of the enzyme specificities Phosphatases hydrolyze phosphate–ester bonds in all types of phosphate esters Chymotrypsin catalyzes the hydrolysis  Stereochemical Specificity –The enzyme can distinguish between stereoisomers –Chirality is inherent in an active site (as amino acids are chiral compounds) L-Amino-acid oxidase catalyzes reactions of L-amino acids but not of D-amino acids SPECIFICITY OF ENZYMES FACTORS AFFECTING ENZYMES  FACTORS AFFECTING ENZYMES Temperature With increased temperature the Enzymes kinetics increases –More collisions –Increased reaction rate FACTORS AFFECTING ENZYMES : pH Most enzymes are active over a very narrow pH range –Protein & amino acids are properly maintained –Small changes in pH (low or high) can result in enzyme denaturation & loss of function Each enzyme has its characteristic pHOPT, which usually falls within  FACTORS AFFECTING ENZYMES : Substrate Concentration If [enzyme] is kept constant & the [substrate] is increased –The reaction rate increases until a saturation point is met At saturation the reaction rate stays the same even if the [substrate] is increased  FACTORS AFFECTING ENZYMES : Enzyme Concentration If the [substrate] is kept constant & the [enzyme] is increased –The reaction rate increases –The greater the [enzyme], the greater the reaction rate  Question time What happens to the enzymes when the body temperature rises from 37ᵒC to 42ᵒC? If an enzyme has broken down and is non- functional, what would happen to the chemical reaction normally facilitated by the enzyme? Explain.  Michaelis-Menten Equation The Michaelis-Menten equation describes the relationship between the rate of an enzyme- catalyzed reaction and the substrate concentration.  Km and Vmax Values Km: Vmax: A measure of an enzyme's The maximum rate of an affinity for its substrate. enzyme-catalyzed A lower Km indicates a reaction. higher affinity for the It is determined by the substrate. enzyme's turnover A higher Km indicates a number (the number of lower affinity for the substrate molecules substrate. converted to product per unit time).  Enzyme Inhibition –A substance that slows down or stops the normal catalytic function of an enzyme by binding to the enzyme Three types of inhibition: Reversible Reversible non- Irreversible competitive competitive inhibition inhibition inhibition Competitive inhibitor LINEWEAVER-BURKE PLOT FOR COMPETITIVE INHIBITION Competitive inhibitors do not Competitive change Vmax of enzyme inhibitor reactions. They increase the apparent Km value. This is because they bind 1 [V] reversibly to the active site. Increasing substrate Uninhibited concentration can overcome enzyme their inhibition. Unchanged Vmax but 1 Vmax increased Km reflect the effects of competitive inhibition. -1 1 Km [S] Competitive inhibitor  LINEWEAVER-BURKE PLOT FOR NONCOMPETITIVE INHIBITION Noncompetitive inhibitor Vmax decreases because a noncompetitive inhibitor reduces the concentration of the enzyme- 1 substrate (ES) complex that can [V] proceed to form products, resulting Uninhibited in differing y-intercepts on enzyme Lineweaver-Burk plots. 1 Km remains unchanged because Vmax the substrate can still bind to the enzyme 1 1 - [S] Km Competitive inhibitor  LINEWEAVER-BURKE PLOT FOR UNCOMPETITIVE INHIBITION Uncompetitive inhibitor Uncompetitive inhibitors decrease both Vmax and 1 Km, hence the [V] production of parallel lines in uninhibited and Uninhibited inhibited reactions. 1 enzyme Vmax 1 1 [S] - Km  Uncompetitive Inhibition The inhibitor binds to the enzyme-substrate complex. Decreases both Vmax and Km. Cannot be reversed by increasing the substrate concentration.  Allosteric site It refers to the regulatory site of an enzyme, which provides a binding site for the effector or non- substrate molecules that either activate or inhibit the enzyme’s catalytic efficiency.  Types of Allosteric Positive Allosteric Regulation: Binding of the effector increases the enzyme's affinity for the substrate, leading to an increase in enzyme activity. Negative Allosteric Regulation: Binding of the effector decreases the enzyme's affinity for the substrate, leading to a decrease in enzyme activity. Drugs Inhibiting Enzyme Activity ACE Inhibitors Inhibit Angiotensin- Converting Enzyme Lowers blood pressure  Drugs Inhibiting Enzyme Activity Penicillin's –β-lactam antibiotics inhibit transpeptidase Transpeptidase enzyme strengthens the cell wall –Without transpeptidase enzyme (inhibited by Penicillin) >>> weakened cell wall, bacteria dies  Medical Uses of Enzymes Lactate dehydrogenase(LDH) is normally not found in high levels in blood, as it is produced in cells –Increased levels of LDH in the blood indicate myocardial infarction (MI) (Heart attack) Tissue plasminogen activator (TPA) activates the enzyme plasminogenthat dissolves blood clots Used in the treatment of MI  I. Enzymes as Therapeutic Agents (Drugs) S. Enzyme Disease/therapy No 1 Streptokinase Clot lysis in myocardial infarction, trauma, bleedings 2 Aspariginase Acute lymphocytic leukemia 3 Adenosine Severe combined immuno- deaminase deficiency syndrome (SCID)  Enzymes as Drug Targets Enzyme targeting Drug Dihydrofolate reductase Antifolates: methrotrexate (cancer) pyrimethamine (protozoa, malaria) Xanthine oxidase Allopurinol (hyperuricemia, gout) (Purine metabolism) Thymidylate synthase 5-Fluorouracil & (Pyrimidine metabolism) 5-fluorodeoxyuridine (cancer) Glycopeptide transpeptidase Antibiotics, penicillin HIV-Reverse transcriptase 3’-azido-2’,3’-dideoxythymidine (AZT) HIV & SARS proteases Ritonavir, saquinavir (clinical trial phase)  II. Enzymes as Diagnostic Agents Enzyme Cause of elevated plasma level Acid phosphatase - ACP Prostatic cancer Alkaline phosphatase – ALP Rickets, hypoparathyroidism, osteomalacia, obstructive jaundice, cancer of bone/liver Alanine aminotransferase – ALT Hepatitis, jaundice, circulatory faillure Aspartate aminotransferase – AST Myocardial infarction, muscle damage, anemia, Amylase - AM Acute pancreatitis, peptic ulcer -Glutamyl transferase – GMT Hepatitis, alcoholic liver damage, cholestasis Creatine kinase – CK Myocardial infarction, Muscular dystrophy Lactate dehydrogenase – LD1 > LD2Myocardial infarction, kidney disease, LD2, LD3 Leukemia LD5 Liver disease, muscle damage  Question time https://www.rgpv.ac.in/campus/PY/enzymes_ ppt.pdf https://nios.ac.in/media/documents/dmlt/Bio chemistry/Lesson-08.pdf https://byjus.com/biology/enzymes/