Enzymes (1) – Comprehensive Study Notes

Introduction to Cellular Catalysis

• Most spontaneous chemical reactions inside a living cell proceed far too slowly to sustain life.
  • A typical uncatalyzed biological reaction can require ≈ 750 000 000 years to reach completion.
• To make metabolism feasible, cells employ biological catalysts called enzymes that dramatically accelerate reaction rates.

General Concepts & Core Definitions

• Catalyst
  • Substance that increases the rate (velocity) of a chemical reaction while emerging unchanged at the end of the process.
  • Does not alter the reaction’s equilibrium constant.
• Reaction rate / velocity
  • Expressed as the change in moles or grams of reactant or product per unit time (e.g., $\text{mol}\,\text{min}^{-1}$).
• Enzymes
  • Primary biological catalysts; the vast majority are proteins, though catalytic RNA molecules (ribozymes) exist.
• Substrate (S)
  • The specific molecule upon which an enzyme acts to produce product (P).

Fundamental Properties of Enzymes

1. Extreme efficiency – can increase reaction rates by factors of $10^{6}$ to $10^{12}$.
2. Lower the activation energy (Eₐ) required to reach the transition state, thereby speeding up reactions.
3. Equilibrium neutral – final equilibrium (Keq) of the reaction remains unchanged.
4. Recovered unchanged – enzyme’s structure and quantity remain the same after catalysis.
5. Possess an active site – specialized pocket/groove that binds substrate.
6. High specificity
   • Substrate specificity (what they bind).
   • Reaction specificity (what transformation they catalyze).
7. May require additional helpers – coenzymes, metal cofactors, or prosthetic groups.
8. Regulatable – activity can be modulated by inhibitors, activators, covalent modification, gene expression, etc.

Activation-Energy Profile

• Uncatalyzed reaction: large activation-energy barrier must be overcome.
• Enzyme-catalyzed reaction: barrier is lowered, transition state stabilized.

graph LR; A[Reactants] -- High Ea --> TS1[Transition State]; TS1 --> B[Products]; A -- Lower Ea\nenzyme--> TS2[Transition State*]; TS2 --> B

• Energy difference illustrated on slides:
  • (a) Non-catalyzed vs. (b) Enzyme-catalyzed.
  • Activation energy ↓, energy released (ΔG) unchanged.

Chemical shorthand for catalytic cycle:
$(1)\;S + E \rightleftharpoons ES \rightarrow EP \rightarrow E + P$

Architecture of the Active Site

• Pocket or groove formed by the enzyme’s tertiary (and sometimes quaternary) structure.
• Composed of binding residues (orient the substrate) and catalytic residues (perform chemistry).
• Provides complementary shape, charge, polarity, and hydrophobic/hydrophilic character to the substrate.

Illustrative cartoon: substrate sits within a shaded “well,” forming the enzyme–substrate (ES) complex before chemistry occurs.

Enzyme Classification (IUBMB)

IUBMB divides all enzymes into 6 major classes, each further subdivided; examples and subclasses included:

1. Oxidoreductases – catalyze redox reactions
   • Subclasses: Oxidases, Dehydrogenases
   • Example: Alcohol dehydrogenase.
2. Transferases – transfer functional groups between molecules
   • Subclasses: Aminotransferases, Kinases (phosphotransferases)
   • Example: Glucokinase, Hexokinase.
3. Hydrolases – catalyze hydrolytic cleavage (addition of water)
   • Example: Peptidases.
4. Lyases – add/remove groups to form double bonds, or remove CO₂/NH₃/H₂O without hydrolysis
   • Example: Pyruvate decarboxylase.
5. Isomerases – intramolecular rearrangements
   • Subclasses: Epimerases, Isomerases.
6. Ligases – join two molecules with concurrent ATP hydrolysis
   • Often termed synthetases.

Nomenclature Rules

• Trivial naming by substrate + “ase”
  • Lipase, Amylase, Protease.
• Naming by reaction type
  • Alcohol dehydrogenase (removes H₂ from alcohol).
• Historical exceptions: Trypsin, Pepsin, Chymotrypsin etc.
• Systematic IUB name = “substrate” + “reaction type” + “ase”
  • e.g., Amino acid oxidase, Lactate dehydrogenase.

EC Numbering for Research

• Four-digit Enzyme Commission (EC) code $a.b.c.d$
  1. Class
  2. Subclass
  3. Sub-subclass
  4. Unique enzyme ID
• Example: Hexokinase → EC 2.7.1.1

Specificity Levels & Mechanistic Models

• Absolute specificity – enzyme acts on only one substrate (e.g., Glucokinase acts only on glucose).
• Broad specificity – enzyme recognizes several structurally related substrates (e.g., Hexokinase phosphorylates glucose, fructose, mannose).

Model 1: Lock-and-Key

• Enzyme’s active site is rigid; substrate must perfectly fit like a key into its lock.
• Explains high specificity but fails to account for conformational flexibility observed experimentally.

Model 2: Induced Fit

• Proposed by Koshland.
• Active site is partially pre-formed; substrate binding induces conformational change, optimizing contact and catalysis.
• Better explains dynamic protein behavior.

Annotated schematic shows:

• “a b c” residues of enzyme adjusting around substrate to form tight ES complex.

Molecular Composition of Enzymes

• Simple (apoenzymes) – purely protein, catalytically inactive without any additional component (if required).
• Complex (holoenzymes) – protein component + non-protein component; fully active form.
  • $\text{Holoenzyme} = \text{Apoenzyme} + \text{Cofactor / Coenzyme / Prosthetic group}$

Non-Protein Components

1. Cofactors – inorganic ions (usually metal ions)
   • Examples: $K^+$, $Na^+$, $Ca^{2+}$, $Mg^{2+}$, $Zn^{2+}$.
2. Coenzymes – small organic molecules derived largely from vitamins; loosely bound
   • Examples: NAD⁺ (vitamin B₃, niacin), FAD (vitamin B₂, riboflavin).
3. Prosthetic groups – organic or inorganic moieties tightly or covalently attached to the enzyme.

Not every enzyme needs these helpers; e.g., digestive proteases pepsin & trypsin function without cofactors.

Regulatory Capacity (mentioned implicitly)

• Enzyme activity can be fine-tuned by:
  • Allosteric effectors.
  • Covalent modification (phosphorylation, acetylation).
  • Proteolytic activation (zymogens → active enzymes).
  • Gene-level control of synthesis/degradation.

Real-World & Physiological Significance

• Understanding enzyme mechanisms underpins drug design, metabolic engineering, and diagnosis of disease (e.g., elevated plasma enzymes as biomarkers).
• Vitamin deficiencies directly impair coenzyme availability → metabolic disorders.
• Industrial biotechnology exploits robust enzymes (e.g., thermostable proteases in detergents).

Numerical & Symbolic Summary

• Uncatalyzed biological reaction time: $\approx 7.5\times10^{8}\ \text{years}$
• Catalytic acceleration: up to $10^{12}\text{-fold}$ faster.
• Generic enzyme equation: $E + S \rightleftharpoons ES \rightarrow E + P$.
• Energy profile (qualitative): $E<em>a^{\text{cat}} < E</em>a^{\text{non-cat}}$ while $\Delta G^{\circ}!!_{\text{reaction}}$ unchanged.