L23: Enzyme Essentials

• What are enzymes classified as?

  Enzymes are catalytic proteins.
• Why is a little bit of chemistry needed to appreciate how enzymes work?

  Because enzymes' speed and efficiency can be expressed numerically, requiring mathematical handling.
• What are co-factors in relation to enzymes?

  Co-factors are small molecules required for enzyme activity that are not part of the enzyme itself.
• Give an example of a co-factor mentioned in the study material.

  The active site zinc ion in human carbonic anhydrase II.
• What are the types of co-factors and their sources?

  • Organic co-factors: derived from vitamins or non-vitamins (e.g., quinone, nucleotides)
  • Inorganic co-factors: metal ions (e.g., zinc, cobalt, copper, iron, magnesium, manganese, molybdenum)
• What is an apoenzyme?

  An apoenzyme is an enzyme lacking an essential co-factor.
• What is a holoenzyme?

  A holoenzyme is the complex formed between an apoenzyme and a co-factor.
• What are the key points about the function of enzymes?

  1. Enzymes increase the rate of chemical reactions.
  2. They increase both forward and reverse rates without changing chemical equilibria.
  3. Enzymes do not undergo net change when acting as catalysts.
• How do enzymes affect the reaction of CO2 and H2O?

  Enzymes like carbonic anhydrase catalyze both forward and reverse rates of the reaction.
• What is the reaction catalyzed by carbonic anhydrase?

  CO2 + H2O ⇌ HCO3- + H+
• What is the turnover number (k_cat) for carbonic anhydrase?

  k_cat = 1,000,000 s^-1
• How can catalytic power be described in relative terms?

  It can be described as acceleration compared to the non-catalyzed rate.
• What is the catalytic constant (k_cat) for catalase?

  k_cat = 10^7 s^-1
• Which enzyme has the lowest catalytic constant listed?

  Lysozyme with k_cat = 0.5 s^-1.
• Under what condition is a catalytic constant considered constant?

  A catalytic constant is constant if conditions do not change.
• How does pH affect the catalytic constant of carbonic anhydrase II?

  Maximal activity occurs at high pH, around pH 9.
• What conditions can alter catalytic constants?

  • pH: Enzymes work efficiently over restricted pH ranges (e.g., alkaline phosphatase at pH 8-10, pepsin at pH 1.5-1.6).
  • Temperature: Different enzymes have optimal temperatures (e.g., Pseudoalteromonas β-galactosidase at 23°C, Taq polymerase at 75-80°C).
• What are the classifications of enzymes according to the Enzyme Commission number (EC)?

  1. EC 1: Oxidoreductases (e.g., lactate dehydrogenase)
  2. EC 2: Transferases (e.g., hexokinase)
  3. EC 3: Hydrolases (e.g., glucose-6-phosphatase)
  4. EC 4: Lyases (e.g., carbonic anhydrase)
  5. EC 5: Isomerases (e.g., triose-phosphate isomerase)
  6. EC 6: Ligases (e.g., amino-acyl tRNA synthase)
• What does it imply if an enzyme can discriminate between similar molecules?

  It implies complementarity between the enzyme’s active site and the substrate.
• What is an example of an enzyme that can differentiate between optical isomers?

  Isocitrate dehydrogenase can differentiate between optical isomers.
• What model did Emil Fischer propose in the 1890s regarding enzyme activity?

  Fischer proposed the 'lock and key' model for enzyme activity.
• What was Daniel Koshland's observation regarding hexokinase?

  Koshland observed that water, an analogue of glucose, does not react with hexokinase.
• What does the 'induced fit' model propose about enzyme activity?

  The 'induced fit' model proposes that substrates cause a change in the 3-D structure of the active site for proper orientation of catalytic groups.
• How did the scientific community initially respond to Koshland's 'induced fit' model?

  The scientific community largely rejected Koshland's ideas, favoring Fischer's 'lock and key' model instead.
• What structural evidence emerged in the 1970s regarding the induced fit model?

  Structural evidence showed that glucose induces enzyme folding in hexokinase.
• What is the active site of an enzyme?

  The active site is the part of the enzyme that is in contact with the substrate and involved in catalyzing the reaction.
• What is the transition state in a chemical reaction?

  The transition state is a high-energy intermediate structure that is neither substrate nor product.
• What happens during the transition state of a chemical reaction?

  Old bonds are weakened and new bonds begin to form during the transition state.
• What model describes how glucose induces enzyme folding in hexokinase?

  Induced fit model
• What is the active site of an enzyme?

  The active site is the part of the enzyme that binds the substrate and catalyzes the reaction.
• What is the transition state in a chemical reaction?

  The transition state is a high-energy intermediate that is neither substrate nor product.
• What is activation energy in a chemical reaction?

  Activation energy is the energy required to reach the transition state.
• How do enzymes enhance the rate of reactions without altering free energy?

  Enzymes lower the activation energy required to reach the transition state.
• What is the catalytic triad in chymotrypsin?

  The catalytic triad consists of three key amino acids that are brought together in the enzyme's tertiary structure.
• What interactions occur between the substrate and the active site during enzyme catalysis?

  • Multiple weak non-covalent interactions
  • Types include electrostatic interactions, hydrogen bonds, and Van der Waals forces
  • These interactions lower the activation energy and facilitate the formation of the transition state
• What type of enzyme is thrombin and what is its function?

  Thrombin is a specific protease that participates in blood coagulation.
• How does chymotrypsin differ from trypsin in terms of substrate specificity?

  Chymotrypsin hydrolyzes peptide bonds at the C-terminal side of aromatic amino acids, while trypsin cleaves at the carboxyl side of positively charged residues.
• What are the steps in the reaction mechanism of chymotrypsin?

  1. Docking of the substrate peptide leads to induced fit.
  2. His57 acts as a base catalyst, activating Ser195.
  3. Ser195 attacks the target peptide bond.
  4. Formation of a tetrahedral intermediate.
  5. Collapse of the intermediate to form an acyl-enzyme.
  6. Nucleophilic attack by water resolves the acyl-enzyme.
  7. Release of the N-terminal fragment and regeneration of the enzyme.
• How does trypsin activate chymotrypsin?

  Trypsin activates chymotrypsin by proteolytic cleavage, which helps form the active site.