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Download Lecture 4 Enzymes Catalytic proteins Enzymes Enzymes Enzymes
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10/21/10 Enzymes Lecture 4 Enzymes • Are a type of protein that acts as a catalyst-speeding up chemical reactions • A catalyst is defined as a chemical agent that changes the rate of a reaction without being consumed by the reaction • Enzymes are biological catalysts • Chemical reactions involve the breaking and forming of bonds-requires energy Catalytic proteins Enzymes Enzymes • Activation energy is the initial investment of energy needed to start a reaction • What enzymes do therefore is: Reactants Transi5on state Products (ready to react) • In most chemical reactions this is provided in the form of heat Enzymes act here • Biochemists call ‘reactants’ substrates Substrate • humans need to maintain a temperature of 37⁰C Transi5on state Products (ready to react) Enzymes act here • Enzymes provide the activation energy for reactions to proceed at normal temperatures in biological systems • Enzymes are substrate-‐specific – each substrate fits into the enzyme’s ac5ve site 1 10/21/10 Eg. Sucrase 1 Active site is available for a molecule of substrate, the reactant on which the enzyme acts. Substrate (sucrose) How do they work? 2 Substrate binds to enzyme. Transi5on state Glucose OH Fructose Enzyme (sucrase) H 2O H O 4 Products are released. 3 Substrate is converted to products. Hydrolysis of sucrose involves the breaking of the bond between glucose and fructose the breaking of a bond in the water molecule the forming of 2 new bonds • Complex molecules have the potential to breakdown spontaneously (2nd law of thermodynamics= the universe tends towards disorder) • They continue to exist because the initial activation energy required to breakdown can’t be reached to initiate a breakdown reaction • An enzyme lowers the activation energy required for a reaction enabling the substrates to absorb enough energy even at a moderate temperature to reach transition state Enzymes “active site” Enzyme shape • Enzymes are substrate specific – The substrate fits like a key in a lock – When the substrate and enzyme are joined the catalytic action of the enzyme converts the substrate to product • The specificity of an enzyme for a substrate results from the shape of the enzyme –due to a compatible fit between the active site and the substrate • The substrate binds to the “active site” of the enzyme -This is the catalytic centre of the enzyme - The active site is typically a pocket or groove on the surface of the enzyme • Is usually formed by only a few amino acids • Is not rigid • The side chains of the aa’s in the enzyme interact with the chemical groups of the substrate- enzyme changes shape and fits even better around the substrate- INDUCED FIT • Induced fit brings the chemical groups of the active site into new positions enhancing the ability of the enzyme to catalyse the reaction • The rest of the protein structure provides the structural framework that determines the configuration of the active site 2 10/21/10 Induced fit Induced fit Induced fit Types of Enzymes 6 major types 3 10/21/10 Types of Enzymes • ATPases - hydrolyse ATP. Many proteins with a wide range of roles have an energy-harnessing ATPase activity as part of their function, for example, motor proteins such as myosin and membrane transport proteins such as the sodium–potassium pump. • Kinases - catalyze the addition of phosphate groups to molecules. Protein kinases are an important group of kinases that attach phosphate groups to proteins. • Nucleases - break down nucleic acids by hydrolyzing bonds between nucleotides. Types of Enzymes • Simple enzymes composed wholly of protein • Complex enzymes-called holoenzymes – Composed of protein (apoenzyme)+ a simple small organic molecule (cofactor or prosthetic group) • Phosphatases - catalyze the hydrolytic removal of a phosphate group • Polymerases - catalyze polymerization reactions such as the synthesis of DNA and RNA. – Metalloenzymes are composed of protein (apoenzyme) + metal • Proteases - break down proteins by hydrolyzing bonds between amino acids. 6 important features of enzymes 1. Side chains of amino acids make up the active site 2. Substrates bind via weak interactions: hydrogen bonds, hydrophobic interactions 3. Act at optimal pH, temperature and substrate concentration 4. May require other factors to work 5. Can be inhibited by drugs 6. Are activated or inhibited by chemical modification Features of enzymes 1. Side chains of amino acids make up the active site – Dependent on R group chemistry – Dependent on final folding 3. Substrates bind via weak interactions: – hydrogen bonds – hydrophobic interactions 4 10/21/10 Features of enzymes 3. Enzymes act at optimal pH and temperatures: Features of enzymes 3. Enzymes act at optimal substrate concentration Michaelis Menton Kinetics Most enzymes are like this Enzyme ac5vity How well the enzyme is working = rate of activity Enzyme ac5vity 37oC 7.4 temp pH Concentration of substrate Excep5ons: Thermophiles 70⁰C Excep5ons: pH 2 in stomach – pepsin pH 8 in intes5ne -‐ trypsin Features of enzymes 3. Enzymes act at optimal substrate concentration Features of enzymes 3. Enzymes act at optimal substrate concentration Michaelis Menton Kinetics Michaelis Menton Kinetics How well the enzyme is working = rate of activity Concentration of substrate Initial rate 5 10/21/10 Features of enzymes Can define how well a substrate binds to an enzyme by the Km value Km value is the substrate concentration at which the enzyme is working at 50% activity Big Km = bad binding Low Km= good binding Features of enzymes 4. Many require non-protein helpers to aid catalysis - ‘co-factors’ Eg: calcium, iron. • Michaelis Menton Kinetics • Features of enzymes 5. Inhibition of enzymes by drugs or molecules in the body • Irreversible-destroys the enzyme • Reversible – Competitive-inhibitor resembles the substrate – Noncompetitive-inhibitor binds to another site on the emzyme called an allosteric site If the co-factor is organic it is a coenzyme - Most vitamins are coenzymes Eg: vitamin C Vitamin C is used by an enzyme to cross-link collagen. Vitamin C deficiency causes scurvy – a disease of dysfunctional cross-linking of collagen Features of enzymes 5. Many drugs inhibit enzymes • Drugs can compete with the substrate and prevent the reaction happening HMG-‐CoA Mevalonic acid Cholesterol ENZYME: HMG-‐CoA reductase Eg. Statins inhibit HMG-Co Reductase, a key enzyme in cholesterol synthesis HMG-‐CoA – statins therefore lower cholesterol and protect or against heart disease. Lovostatin competes with the substrate for the active site HMG-CoA reductase lovosta5n 6 10/21/10 Features of enzymes Features of enzymes 6. Cellular enzymes can be activated by “phosphorylation” 6. Are activated or inhibited by a chemical modification • Other enzymes add the phosphate group to the other enzyme, -these enzymes are called kinases – Phosphorylation –addition of phosphate grp • Kinases are said to “phosphorylate” the protein – Acetylation –addition of an acetyl group • Eg: Adrenalin activates Phosphorylase kinase which phosphorylates Glycogen Phosphorylase. -This enzyme in turn breaks down glycogen to release glucose – Dephosphorylation-removal of a phosphate group – Deacetylation-removal of an acetyl group Signal cascade In summary... Receptor protein Adrenaline (hormone protein) signals Phosphorylase Kinase (protein kinase enzyme) P GP GP Glycogen Phosphorylase Glycogen glucose P =Phosphate group • Enzymes are catalytic proteins • Lower the initial activation energy required for a reaction to happen • Are specific for their substrate – shape • The active site is usually made up of only a few amino acid side chains • Induced fit • There are 6 important features Adrenaline results in the release of glucose which allows for rapid produc4on of energy 7