3) Enzymes

• Enzymes


  • They are globular proteins with a specific 3D structure determined by their sequence of amino acids
  • The hydrophilic R groups of amino acids are towards the outside, making them water-soluble
  • Enzymes may show tertiary or quaternary levels of protein structures
  • Enzymes act as biological catalysts, speeding up chemical reactions without being changed or used
  • Enzymes have a cleft or depression called an active site formed of certain amino acids in the polypeptide chain, and they are specific because the shape of the active site is complementary to only 1 type of substrate
• Enzyme Properties


  Enzymes are synthesized on the ribosomes, modified in the RER and Golgi to be used intracellularly or extracellularly
• Lock & Key Hypothesis

  The substrate acts as a key and the enzyme's active site acts as a lock. During catalysis, the substrate fits inside the active site, forming the enzyme-substrate complex, and then products are formed and leave the active site
• Induced Fit Hypothesis


  The enzyme's active site does not "fit" the substrate until the substrate actually enters the site. When the substrate starts binding, it induces a conformational change in the active site shape, molding around the substrate to fit perfectly and become complementary to form the enzyme-substrate complex
• Mode of action


  The temporary hydrogen bonds between specific R groups of the enzyme and the substrate hold the substrate in a particular way inside the active site, bringing reactive groups close together, placing a strain on the bonds, and facilitating Redox reactions to transfer electrons, thus reducing the activation energy of the reaction
• Rate of Reaction


  The progress of an enzyme-catalyzed reaction can be followed by measuring the amount of products produced per unit time or the amount of substrate used per unit time
• Enzymes lower the amount of activation energy needed for reactions, often making reactions happen at a lower temperature than they could without an enzyme
• Activation energy is the energy needed by reactant molecules to reach an unstable transition state to be converted into products, often provided as heat
• The reaction rate slows down as more substrate is converted into product until eventually all substrates are converted into products and the reaction stops
• As more substrate is converted into product


  Fewer substrates are available to bind with enzymes, thus the rate of the reaction slows down until eventually all substrates are converted into products and the reaction stops
• Initial rate of reaction

  The rate at the start of the reaction
• Measuring the initial rate of reaction

  1. Calculating the slope of a tangent to the curve as close to time 0 as possible

  2. Reading off the graph the amount of oxygen given off in the first 30 seconds
• The rate of oxygen production in the first 30 seconds is 2.7 cm3 of oxygen per 30 seconds, or 5.4 cm3 per minute
• Method 1 to measure the rate of the reaction involving Starch + water to Maltose
• Method 1 to measure the rate of the reaction

  Measure the rate at which starch disappears from the reaction mixture by taking samples at known times and adding each sample to iodine in potassium iodide solution. Starch forms a blue-black colour with this solution. Use a colorimeter to measure the intensity of the blue-black colour obtained and calculate the initial reaction rate from the slope
• Method 2 to measure the rate of the reaction involving Starch + water to Maltose
• Method 2 to measure the rate of the reaction

  Mix starch, iodine in potassium iodide solution, and amylase in a tube, and take regular readings of the colour of the mixture in one tube in a colorimeter. Note that iodine interferes with the rate of the reaction and slows it down
• The rate of reaction can be calculated by the equation ROR = 1 / time taken
• As temperature increases, the rate of reaction increases till it reaches a peak at 40°C, which is the optimum temperature for that enzyme. Further increase in temperature results in a steep decrease of the rate of reaction to reach zero at 55°C
• Factors affecting Rate of Enzyme Action: Temperature
• PH of the solution is the measure of its hydrogen ion concentration. The lower the pH, the higher the hydrogen ion concentration
• The rate of enzyme action is highest at pH 7, which is the optimum pH. An increase or decrease from pH 7 results in a decrease in the rate of reaction, till the rate of reaction reaches zero at pH 3 and pH 11
• Each enzyme has an optimum pH at which it works best. Some enzymes work well in acidic pH while others work well in alkaline. Most enzymes work best at a neutral pH of 7
• Tertiary structure changes along with the active site shape


  Preventing the binding of the substrate to it
• Each enzyme has an optimum pH at which it works best
• Enzymes working well at different pH levels

  • Some work well in acidic pH
  • Others work well in alkaline pH
  • Most work best at a neutral pH of 7
• The more enzymes present, the more active sites will be available for the substrates to bind into and form ES complexes
• At low enzyme concentration


  Enzyme concentration is the limiting factor for the reaction
• At high enzyme concentrations


  Adding more enzyme has no effect on the reaction as enzyme concentration is no longer limiting while substrate concentration is
• Because the quantity of hydrogen peroxide is the same in all five reactions, the total amount of oxygen eventually produced will be the same; so, if the investigation goes on long enough, all the curves will meet
• As substrate concentration increases, the rate of reaction increases till it reaches a plateau
• Increasing substrate concentration


  Increases the number of collisions between substrate & enzyme forming more ES complexes per unit time thus increasing rate of reaction
• At high substrate concentration

  All enzyme active sites are saturated and the enzyme is working at its maximum rate (Vmax) so the enzyme concentration becomes the limiting factor
• Km


  A measure of the affinity of the enzyme for its substrate
• Km is the point where half the active sites of the enzyme are occupied by substrate
• The higher the affinity of the enzyme for the substrate, the lower the substrate concentration needed for this to happen
• The higher the affinity, the lower the Km and the quicker the reaction will proceed to its maximum rate
• Turnover rate is the number of substrate molecules converted into products by one enzyme molecule per unit time
• Vmax is the maximum rate of reaction when the enzyme is fully saturated
• Michaelis Menten constant (Km) is the substrate concentration at half the Vmax, it’s a measure of enzyme affinity to substrate