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Enzymes - Study Notes

Objectives

  1. To explain that enzymes are globular proteins that catalyse metabolic reactions.
  2. To define metabolism, anabolism, and catabolism.
  3. To explain the mode of action of enzymes in terms of:
       - Active site
       - Enzyme and/or substrate complex
       - Lowering of activation energy
       - Enzyme specificity.
  4. To explain the properties of enzymes.
  5. To explain the induced-fit hypothesis.
  6. To explain the effects of:
       - pH
       - Temperature
       - Enzyme concentration
       - Substrate concentration on enzyme action.
  7. To construct and interpret graphs showing the effects of the above factors on enzyme action.
  8. To explain the effects of competitive and non-competitive inhibitors on enzyme activity.
  9. To use succinic dehydrogenase, nicotine, and insecticides (pyrethroids) as examples of enzyme inhibitors.
  10. To investigate the effects of temperature and substrate concentration on enzyme-catalysed reactions, and explain these effects.

Introduction

  • Biological Catalysts: Enzymes are known as biological catalysts that facilitate chemical reactions in living organisms.
  • Structure: Mostly globular proteins with specific tertiary shapes, which can also include non-protein catalysts such as ribozymes (RNA molecules) and abzymes (antibodies).
  • Function: They increase the rate of metabolic reactions by lowering the activation energy and are usually specific to only one reaction.

What are Globular Proteins?

  • 3D Structure: Globular proteins exhibit three-dimensional ball-like structures where hydrophobic regions are internal and hydrophilic regions are external, making them water-soluble.
  • Roles: Typically, they perform metabolic roles, including enzymes in organisms and plasma proteins and antibodies in mammals.

Nomenclature of Enzymes

  • Classification System: Governed by the International Union for Biochemical Societies (IUBMB).
  • Naming Convention: An enzyme’s name indicates the substrate it acts upon and the class of enzyme, usually ending with the suffix –ase, e.g., maltase, sucrase, and amylase.

Naming Conventions

  • Based on Reactions: Enzymes are named after the reactions they catalyse.
      - Examples:
        - Sucrase (breaks down sucrose)
        - Protease (breaks down proteins)
        - Lipase (breaks down lipids)
        - DNA polymerase (builds DNA by adding nucleotides).

Classification of Enzymes by Function

There are several classes of enzymes based on their function:

  1. Oxidoreductases: Transfer hydrogen atoms, electrons, or oxygen atoms from substrates to acceptor molecules (e.g., oxidases, dehydrogenases).
  2. Transferases: Transfer small groups of atoms from one substrate to another (e.g., transaminases).
  3. Hydrolases: Split chemical bonds through hydrolysis (e.g., lipases that work on ester linkages in lipids).
  4. Isomerases: Catalyse internal rearrangements of atoms within a substrate (e.g., phosphoglucomutase).

Metabolism

  • Definition: Metabolism is the sum of chemical reactions occurring within each cell of a living organism, providing energy for vital processes and for synthesising new organic materials.
  • Role of Enzymes: Enzymes control metabolic pathways by changing the substrate at each step to achieve the final product.

Anabolic and Catabolic Pathways

  1. Anabolic Pathways: Require energy to build large molecules from smaller ones (biosynthesis).
        - Example: Synthesis of glycogen from glucose.
  2. Catabolic Pathways: Release energy by breaking down large molecules into smaller ones (degradation).
        - Example: Digestion of food where enzymes break down food into absorbable units.
  • Some metabolic pathways can be reversible or irreversible; most pathways are reversible.

Enzymes – How They Function

  • Enzymes operate at lower temperatures compared to the high temperatures that would typically initiate reactions, which would be harmful to cells.
  • Activation Energy: The energy needed to initiate a chemical reaction. Enzymes lower this activation energy, facilitating the reaction.

Activation Energy

  • Definition: Activation energy is the necessary energy required to start a reaction.
  • Function of Enzymes: Enzymes reduce the activation energy requirements for reactions, speeding up the rate without changing the free energy balance between products and reactants.

Energy Profile of Reactions

  • Catalysed vs Uncatalysed Reactions: An uncatalysed reaction requires higher activation energy than a catalysed one. Enzymes create a more favorable energetic environment for the reaction, minimizing the energy barrier.
  • The Gibbs Free Energy remains unchanged between catalysed and uncatalysed reactions.

Active Site

  • Definition: The active site is a sequence of amino acids (3–12) positioned typically on the protein's surface, allowing it to interact easily with substrates.
  • Characteristics: The active site has specific chemical groups and bonds that allow only a complementary substrate to bind, determined by the protein's overall structure.

Substrate

  • Definition: The substrate is the molecule that binds to the active site of an enzyme, having a complementary shape, size, and charge. It is typically smaller than the enzyme.

Lock-and-Key Hypothesis

  • Concept: This model suggests a rigid active site shape that exactly fits the substrate (key) like a lock.
  • Implications: This model highlights enzyme specificity but does not account for flexibility in all enzymes.

Induced-Fit Hypothesis

  • Overview: This more recent model posits that the active site adapts in shape to better accommodate the substrate.
  • Flexibility: Substrates induce a conformational change that leads to a perfect fit and enhanced catalysis, providing a broader range of substrate specificity.

Enzyme Mechanism

  • Principles of Action:
       - Enzymes bring substrates close together in correct orientation.
       - They weaken and break bonds to facilitate the reaction.
       - They lower the energy needed for the reaction and release products after it occurs.
       - They remain unchanged and can be reused for multiple reactions.

Catalytic Cycle of an Enzyme (Example: Sucrase)

  1. Enzyme available with an empty active site.
  2. Substrate (sucrose) binds to the enzyme (sucrose-sucrase complex).
  3. Enzyme converts substrate into products inside the active site.
  4. Products (glucose and fructose) are released, and the active site is available for another substrate.

Features of Enzymes

  1. Recyclable: Enzymes are not consumed in the reactions they catalyse and can be reused.
  2. Require Cofactors: Some enzymes need additional non-protein components for activity such as:
       - Inorganic Ions: E.g., Zn²⁺, Fe²⁺, Mg²⁺, necessary for compatible shapes.
       - Prosthetic Groups: E.g., FAD, permanently attached to enzymes.
       - Coenzymes: Organic substrates that temporarily assist enzymatic activity, e.g., NAD, ATP.
  3. Reversible Reactions: Enzymes can facilitate reactions in either direction depending on concentration gradients.

Inhibition of Enzymes

  • Definition: Inhibition refers to molecules that reduce enzyme activity.
  1. Competitive Inhibitors: Similar to the substrate and bind to the active site, preventing the actual substrate from binding (e.g., penicillin).
  2. Non-Competitive Inhibitors: Bind to sites other than the active site, altering its shape so that the substrate cannot bind (e.g., cyanide).
  3. Allosteric Regulation: Molecules that bind to allosteric sites regulate enzyme activity, altering the active site reversibly (activators enhance the fit, inhibitors obstruct it).

Factors Affecting Rate of Enzyme-Catalysed Reactions

The major factors include:

  1. Substrate Concentration: Increases reaction rate until a saturation point, where all active sites are occupied.
  2. Enzyme Concentration: Increasing enzyme concentration raises reaction rates until substrate is limited.
  3. Temperature: Optimal range enhances activity, high temperatures cause denaturation.
  4. pH: Extreme pH levels can denature enzymes, affecting their efficiency.

Temperature Effects on Enzymes

  • Enzymes are denatured at high temperatures; at extremes (0°C and 100°C), activity is negligible.
  • For every 10°C increase, reaction rates double until optimal conditions are reached (around 40°C) before rate declines due to denaturation.

pH Effects on Enzymes

  • Alterations in pH levels impact enzymes by changing the ionization states of amino acids, which can lead to denaturation at extreme pH levels. Optimal pH varies for each enzyme.

Summary

  • Enzymes are biological catalysts crucial for speeding up chemical reactions by lowering activation energy. They possess an active site that specifically binds substrates, allowing reactions to occur efficiently. Enzymes are highly specific facilitators of biochemical processes in living cells, significantly impacting metabolic activities.