Lab 7/EXP7. Factors Affecting Enzyme Activity PDF

Lab 7/EXP7. Factors Affecting Enzyme Activity  Enzymes speed up metabolic reactions by lowering energy barriers A catalyst is a chemical agent that speeds up a reaction without being consumed by the reaction An enzyme is a catalytic protein For example, sucrase is a...

Lab 7/EXP7. Factors Affecting Enzyme Activity  Enzymes speed up metabolic reactions by lowering energy barriers A catalyst is a chemical agent that speeds up a reaction without being consumed by the reaction An enzyme is a catalytic protein For example, sucrase is an enzyme that catalyzes the hydrolysis of sucrose. Sucrase H2 O O H O H O Sucrose Glucose Fructose (C12H22O (C6H12O (C6H12O 11) 6) 6) © 2018 Pearson Education Ltd.  The Activation Energy Barrier Every chemical reaction between molecules involves bond breaking and bond forming. Loading… The initial energy needed to start a chemical reaction is called the free energy of activation, or activation energy (EA). Activation energy is often supplied in the form of thermal energy that the reactant molecules absorb from their surroundings. © 2018 Pearson Education Ltd. Figure 8.14 The effect of an enzyme on activation energy Course of reaction EA without without enzyme enzym EA with e enzyme is lower Fr Reactants e e Course of ∆G is unaffected e reaction by enzyme n with enzyme er g y Products Progress of the reaction © 2018 Pearson Education Ltd.  How Enzymes Speed Up Reactions In catalysis, enzymes or other catalysts speed up specific Loading… reactions by lowering the EA barrier. Enzymes do not affect the change in free energy (ΔG); instead, they hasten reactions that would occur eventually. © 2018 Pearson Education Ltd. Figure 8.15 Induced fit between an enzyme and its substrate Substrate Active site Enzyme-substrate Enzym complex e © 2018 Pearson Education Ltd.  Substrate Specificity of Enzymes The reactant that an enzyme acts on is called the enzyme’s substrate. tyof Enzymes The enzyme binds to its substrate, forming an enzyme- at an enzyme acts on is called the enzyme s substrate. ds to substrate complex. its substrate, forming an enzyme- substrate e activity of the enzyme converts substrate to product While bound, the activity of the enzyme converts substrate to product. © 2018 Pearson Education Ltd. ’  The reaction catalyzed by each enzyme is very specific. The active site is the region on the enzyme where the substrate binds. Induced fit of a substrate brings chemical groups of the active site into positions that enhance their ability to catalyze the reaction. © 2018 Pearson Education Ltd.  Catalysis in the Enzyme’s Active Site In an enzymatic reaction, the substrate binds to the active site of the enzyme. Enzymes are extremely fast acting and emerge from reactions in their original form. Very small amounts of enzyme can have huge metabolic effects because they are used repeatedly in catalytic cycles. © 2018 Pearson Education Ltd.  The active site and catalytic cycle of an enzyme © 2014 Pearson Education, Inc.  Enzyme Kinetics Vmax is the maximum rate of an enzyme catalyzed reaction i.e. when the enzyme is saturated by the substrate. Loading… Km is the concentration of substrate which permits the enzyme to achieve half Vmax (how easily the enzyme can be saturated by the substrate ) represents substrate affinity. ‫ٮ&واٮ"ﺖ‬ Vmax & Km are constants for enzymes at specific pH and Temperature.  Enzyme Nomenclature Adding the suffix -ase to the name of the substrate that enzymes modify © 2018 Pearson Education Ltd. Effects of Local Conditions on Enzyme Activity An enzyme’s activity can be affected by: general environmental factors, such as temperature and pH. chemicals that specifically influence the enzyme. © 2018 Pearson Education Ltd.  Effects of Temperature and pH Each enzyme has an optimal temperature in which it can function. Each enzyme has an optimal pH in which it can function. Optimal conditions favor the most active shape for the enzyme molecule. © 2018 Pearson Education Ltd. Figure 8.17b Temperature Optimal temperature for Optimal temperature for typical human enzyme (37ºC) enzyme of thermophilic (heat-loving) bacteria (75ºC) R at e of re a ct io n 0 2 4 6 8 100 120 0 0 0 Temperature 0 (ºC) (a) Optimal temperature for two enzymes © 2018 Pearson Education Ltd. Figure 8.17c PH Pepsin (stomach Trypsin (intestinal enzyme) enzyme) R at e of re a ct io n 0 1 2 5 3 4 6 7 8 9 10 pH (b) Optimal pH for two enzymes © 2018 Pearson Education Ltd.  [Substrate] Increase substrate concentration will slow down the rate of the reaction due to enzymatic saturation  Clinical Significance Hyperthermia : degradation of liver enzyme activity liver Failure. H.Pylori infection Damaging effect on acids and pepsin production in the stomach. Paracetamol Overdose saturation of liver enzymes (ex. Glucuronidase) liver toxicity and failure.  Catalase Enzyme Catalase is a common enzyme found in nearly all living organisms exposed to oxygen (such as bacteria, plants, and animals) which catalyzes the decomposition of hydrogen peroxide (H2O2) to water and oxygen (H20 + O2) It is a very important enzyme in protecting the cell from oxidative damage by reactive oxygen species (ROS).  Hydrogen peroxide is a strong oxidizing agent, which tends to disrupt the delicate balance of cell chemistry. too much hydrogen peroxide can lead to cell death. Catalase is located in all major sites of H2O2 production in the cellular environment (such as peroxisomes, mitochondria, cytosol and chloroplast) of higher plants. Potatoes, particularly, contain high amounts of catalase.  In today’s lab we will study: Factors affecting catalase activity [Enzyme] [substrate] Temperature pH  Effect of enzyme concentration Tube Enzym Distilled water H2O2 e (ml) (ml) extra ct (ml) A 0 2 2 B 1 1 2 C Mix all tubes2 and record your 0 2 observation Increasing [enzyme] will increase reaction rate Effect of substrate concentration Tube [H2O2] Distilled Enzyme % water extract (ml) (ml) A 0 2 2 Tube [H2O2] H2O2 Enzyme % (ml) extract (ml) B 0.1% 2 2 C 1% 2 2 D 10% 2 2 Mix all tubes and record your Increasing [substrate] will increase reaction rate to a certain limit because at certain point all enzymes will be occupied (saturated) with substrate molecules  Effect of temperature Tub Enzyme H2O2 Temperatur e extrac (ml) e t (ml) (ºC) A 2 2 4 B 2 2 37 C 2 2 98 Leave all tubes at the specific temperatures for 5 min. Add substrate and leave all tubes for further 10 min at the specific temperatures. Record your observation. Optimum Temperature Each enzyme has a specific temperature at which it performs maximally. low temperatures will not affect enzyme structure however, their activity will be reduced. High temperatures will change the 3D conformation of the enzyme causing changes (degradation) of the enzyme this will result in loosing its function.  High 37C Cold Loading… More bubbles (more Oxygen)  Effect of pH Tube H2O H2O2 Enzyme extract 2 Volum (ml) pH e (ml) A 5 2 2 B 7 2 2 C 9 2 2 Mix all tubes and record your observation  Optimum pH Each enzyme has a specific pH at which it performs maximally. pH that is di pH that is different from the optimal might cause changes in the protein 3D conformation of the enzymes which is the reason standing behind their low activity. Enzyme Optimum pH Salivary amylase 6.8 Stomach protease (pepsin) 1.5–2.0 Pancreatic protease (trypsin) 7.5–8.0