Enzyme Notes

Enzyme Types and Reactions

  • The initial question involves identifying the enzyme type needed for a specific reaction. The reaction shows a peptide bond being broken with the addition of water, resulting in two separate amino acids.

  • Based on the reaction, the correct enzyme type is protease, which catalyzes the hydrolysis of peptide bonds.

Enzyme Models and Function

  • Enzymes function by binding substrates to their active sites, facilitating chemical reactions through proper alignment and bond manipulation.

  • Effective active sites lower the energy threshold required for reactions to occur.

  • Intermolecular forces (IMF) govern substrate binding to the active site, creating an enzyme-substrate complex held together by these forces.

Transition State and Energy Diagrams

  • The transition state complex represents the highest energy formation during an enzyme-catalyzed reaction, leading to product formation.

  • After the reaction, the enzyme-product complex releases the products and regenerates the enzyme.

  • Energy diagrams illustrate catalyzed reactions as multi-step processes with lower activation energy compared to uncatalyzed reactions.

Active Sites and Specificity

  • Enzymes have specific functional groups within their active sites that participate in reactions, requiring precise alignments and energy thresholds.

  • Enzymes are highly specific for their substrates due to the arrangement of amino acid side chains (R groups) within the active site.

  • An example is given of an endoprotease that cleaves peptide bonds after glutamic acid. The active site of such an endoprotease likely contains glycine, serine, aspartic acid, or lysine residues.

Models of Enzyme Action

  • Lock and Key Model: This model suggests a rigid substrate binding to a rigid enzyme, similar to a lock fitting a key.

  • Induced Fit Model: A more dynamic model where the active site is flexible enough to adapt to the shape of the substrate. The enzyme and substrate work together to achieve optimal binding and catalysis.

Enzyme Specificity

  • Absolute Specificity: The enzyme acts on only one substrate. Example: sucrase acts only on sucrose, breaking it down into glucose and fructose.

  • Group Specificity: The enzyme acts only on molecules with specific functional groups. Example: Alcohol dehydrogenase acts on methanol and ethanol.

  • Linkage Specificity: The enzyme acts on a specific type of chemical bond. Example: Pepsin cleaves peptide bonds.

  • Stereochemical Specificity: The enzyme acts on one isomer over another. Example: L-amino acids over D-amino acids.

Enzyme Activity

  • Enzyme activity measures the rate at which an enzyme converts a substrate into a product.

  • Factors affecting enzyme activity:

    • Temperature

    • Substrate concentration

    • pH

    • Enzyme concentration

Effect of Temperature on Enzyme Activity

  • Increasing temperature increases the kinetic energy of molecules, causing them to move faster.

  • Enzymatic reactions generally proceed faster with increasing temperature until an optimal point.

  • High temperatures denature enzymes, causing them to lose their structure and activity. Sterilization using autoclaves denatures proteins.

Optimal Temperature

  • Optimal temperature is the temperature at which an enzyme exhibits maximum activity. For humans, this is around 37^\circ C. Many human enzymes have optimal activity around 40^\circ C before denaturation occurs.

Effect of pH on Enzyme Activity

  • Changes in pH (acidity or basicity) affect the ionization of amino acid side chains, which can alter enzyme structure and activity.

  • Small changes in pH (e.g., 1 pH unit) can significantly change catalytic activity.

  • Enzymes have an optimal pH range (e.g., 7.0-7.5) at which they exhibit maximum activity.

  • Buffers help regulate pH, maintaining optimal conditions for enzyme activity.

Optimum pH for Enzymes

  • Pepsin: Found in the stomach, with an optimal pH of 1.5-2.0, acting on peptide bonds.

  • Lactase: Found in the small intestine, with an optimal pH of 6.0, acting on lactose.

  • Amylase: Found in the pancreas, with an optimal pH of 6.7-7.0, acting on amylose.

  • Trypsin: Found in the small intestine, with an optimal pH of 7.7-8.0, acting on peptide bonds.

  • Lipase: Found in the pancreas, with an optimal pH of 8.0, acting on lipid ester bonds.

  • Arginase: Found in the liver, with an optimal pH of 9.7, acting on arginine.

Substrate Concentration and Enzyme Activity

  • As substrate concentration increases, enzyme activity initially increases.

  • The saturation curve shows that at a certain point, the enzyme is working at its maximum capability, and further increases in substrate concentration do not increase activity.

  • At saturation, all enzyme active sites are occupied, and substrates are waiting for their turn to be processed.

Turnover Number (TON)

  • Turnover number represents the number of substrate molecules transformed per minute per enzyme molecule.

  • TON = \frac{\text{# of substrates transformed}}{\text{minute} \cdot \text{# of enzyme molecules}}

  • Examples of turnover numbers:

    • Carbonic anhydrase: 36,000,000 (CO₂ + H₂O ⇌ H₂CO₃)

    • Catalase: 5,600,000 (2H₂O₂ ⇌ 2H₂O + O₂)

    • Cholinesterase: 1,500,000 (hydrolysis of acetylcholine)

    • Penicillinase: 120,000 (hydrolysis of penicillin)

    • Lactate dehydrogenase: 60,000 (conversion of pyruvate to lactate)

    • DNA polymerase I: 900 (addition of nucleotides to DNA chains)

Enzyme Concentration and Reaction Rate

  • In cells, enzyme concentration is usually lower than substrate concentration.

  • Reaction rate is directly proportional to enzyme concentration: \text{rate} \propto \text{[enzyme]}

Enzymes in Extreme Conditions

  • Extremozymes are enzymes from microorganisms that survive in extreme conditions (e.g., high salt content, pH, pressure, temperature).

  • Standard enzymes may not function optimally under such conditions.

  • Extremozymes are used in enzymatic cleaners that function effectively in cold water.

Enzyme Inhibition

  • Enzyme inhibition refers to the loss of catalytic activity by preventing substrates from fitting into the active site.

  • An inhibitor is a substance that slows or stops catalytic activity by binding to the enzyme.

  • Types of inhibition:

    • Reversible (competitive and noncompetitive)

    • Irreversible

Competitive Inhibition

  • In competitive inhibition, the inhibitor's structure resembles the substrate, allowing it to bind to the active site.

  • The inhibitor competes with the substrate for the active site, blocking substrate binding.

  • Example: Malonate inhibits succinate dehydrogenase by competing with succinate.

Reversible Inhibition

  • Reversible inhibitors bind through weak intermolecular forces, briefly blocking substrate activity.

  • Example: Antihistamines compete with histamine for binding to histamine receptors, alleviating allergy symptoms.

Noncompetitive Inhibition

  • In noncompetitive inhibition, the inhibitor binds to a site other than the active site, altering the enzyme's shape and preventing catalysis.

  • Heavy metals (e.g., Pb^{2+}, Hg^{2+}, Ag^+) are examples of noncompetitive inhibitors.

Competitive vs. Noncompetitive Inhibition

  • Competitive Inhibition: The maximum reaction rate can be achieved with sufficiently high substrate concentrations, overcoming the inhibitor.

  • Noncompetitive Inhibition: The maximum reaction rate is lowered because the enzyme's catalytic activity is reduced, regardless of substrate concentration.

Irreversible Inhibition

  • Irreversible inhibitors form strong covalent bonds with amino acids in the active site, permanently deactivating the enzyme.

  • The inhibitor's structure does not need to resemble the substrate.

Penicillins as Irreversible Inhibitors

  • Penicillins are antibiotics that act as irreversible inhibitors of bacterial enzymes involved in cell wall synthesis.

  • Penicillin contains a \beta-lactam ring that forms a covalent bond with a serine residue in the active site of the bacterial enzyme, permanently inactivating it.

Enzyme Regulation

  • Enzyme regulation turns enzymes on or off to avoid wasting energy.

  • Regulation ensures that enzyme production is adjusted to meet the cell's needs.

Allosteric Enzymes

  • Allosteric enzymes bind a regulator molecule, which may also be a substrate, at a site distinct from the active site.

  • Allosteric enzymes have a quaternary structure with multiple subunits and binding sites.

  • The binding of a regulator molecule to the regulatory site affects the shape and activity of the active site.

Positive and Negative Regulators

  • Positive Regulator (Activator): Enhances substrate binding and increases enzyme activity.

  • Negative Regulator (Inhibitor): Slows or stops substrate binding and decreases enzyme activity.

Feedback Control

  • Feedback control is a regulatory mechanism where the product of a reaction sequence inhibits an enzyme earlier in the sequence.

  • This prevents overproduction of the product.

Proteolytic Enzymes and Zymogens

  • Proteolytic enzymes (proteases) catalyze the cleavage of peptide bonds.

  • Zymogens are inactive forms of proteolytic enzymes.

  • Zymogen activation involves a conformational change that converts the inactive form to the active form.

  • Zymogen names often have the suffix "-ogen" or the prefix "pro-" or "pre-" (e.g., pepsinogen → pepsin).

Covalent Modification

  • Covalent modification activates or deactivates enzymes by adding or removing covalently bound groups.

  • Phosphorylation: Addition of a phosphate group.

  • Dephosphorylation: Removal of a phosphate group.

ATP \xrightarrow{Kinase} ADP + Phosphorylated Protein

Phosphorylated Protein \xrightarrow{Phosphatase} Protein + P_i

Alcohol Groups in Amino Acids

  • Serine, tyrosine, and threonine have hydroxyl (-OH) groups in common, which can react with kinases and phosphatases for phosphorylation/dephosphorylation.

Vitamins

  • Vitamins are organic compounds essential in small amounts.

  • They must be obtained from the diet.

Types of Vitamins

  • Water-Soluble Vitamins: Vitamins C and B; excess is typically flushed out.

  • Fat-Soluble Vitamins: Vitamins A, K, E, and D; can be toxic if taken in excess.

Vitamin Classifications

  • Water-Soluble: Includes ascorbic acid (vitamin C) and B-complex vitamins (thiamine, riboflavin, niacin, pantothenic acid, vitamin B6, biotin, folate, vitamin B12).

  • Fat-Soluble: Includes vitamin A (retinol, \beta -carotenes), vitamin D (cholecalciferol), vitamin K (phylloquinones, menaquinones), and vitamin E (tocopherols).

Vitamin C (Ascorbic Acid)

  • Water-soluble vitamin.

  • Important for blood vessel health.

  • Deficiency leads to scurvy.

Vitamin B

  • Water-soluble.

  • Precursors to coenzymes.

Cofactors and Coenzymes

  • Apoenzyme: An inactive enzyme that becomes active upon binding a coenzyme or cofactor.

  • Holoenzyme: The active enzyme formed when a cofactor or coenzyme binds to the enzyme's active site.

  • Cofactor: A non-protein chemical compound that binds to an enzyme to assist in catalysis. Inorganic cofactors include metal ions (e.g., Mg^{2+}, Fe^{2+}).

  • Coenzyme: A specific type of cofactor that is organic, often including vitamins or derivatives (e.g., NAD^+). Coenzymes bind loosely to enzymes and are typically altered during the reaction.

Functions of Cofactors and Coenzymes

  • Cofactors often play a structural or catalytic role and are not consumed in the reaction.

  • Coenzymes transfer chemical groups from one substrate to another and are typically altered during reactions.

Fat-Soluble Vitamins

  • Vitamin A

  • Vitamin E

  • Vitamin D

  • Vitamin K