Chapter 8: Enzymes – Biological Catalysts

Enzymes: Biological Catalysts

What is a Catalyst?

Catalysts are substances that increase the rate of chemical reactions without being consumed or permanently altered in the process. In biological systems, enzymes serve as highly efficient and specific catalysts.

• Definition: A catalyst increases the rate or velocity of a chemical reaction without being changed in the overall process.

• Thermodynamics: Catalysts accelerate the approach to equilibrium but do not alter the thermodynamic favorability (ΔG) of the reaction.

• Activation Energy: Catalysts lower the activation energy (energy barrier) required to reach the transition state, facilitating the conversion of substrate to product.

Major Classes of Enzymes

Enzymes are classified based on the types of reactions they catalyze. The six major classes are summarized below.

Class	Example (Reaction Type)	Reaction Catalyzed
1. Oxidoreductases	Alcohol dehydrogenase (oxidation with NAD+)	Ethanol → Acetaldehyde
2. Transferases	Hexokinase (phosphorylation)	Glucose → Glucose-6-phosphate
3. Hydrolases	Carboxypeptidase (peptide bond cleavage)	Polypeptide → Peptides/Amino acids
4. Lyases	Pyruvate decarboxylase (decarboxylation)	Pyruvate → Acetaldehyde + CO2
5. Isomerases	Malate isomerase (cis-trans isomerization)	Malate ↔ Fumarate
6. Ligases	Pyruvate carboxylase (carboxylation)	Pyruvate → Oxaloacetate

Additional info: The EC (Enzyme Commission) system is used to classify enzymes based on the reactions they catalyze.

How Enzymes Act as Catalysts: Principles and Examples

Models for Substrate Binding and Catalysis

• Lock-and-Key Model: The enzyme's active site is complementary in shape to the substrate, allowing specific binding without structural change.

• Induced Fit Model: The enzyme undergoes a conformational change upon substrate binding, optimizing the interaction and facilitating catalysis.

• Example: Hexokinase undergoes a significant conformational change when binding glucose, illustrating the induced fit model.

Mechanisms for Achieving Rate Acceleration

Enzymes employ several strategies to accelerate reaction rates:

• 1) General acid/base catalysis (GABC): Enzyme side chains donate or accept protons to stabilize intermediates.

• 2) Covalent catalysis: Formation of a transient covalent bond between enzyme and substrate.

• 3) Electrostatic stabilization: Stabilization of charged transition states by charged or polar residues.

• 4) Proximity effects: Bringing substrates into close proximity and correct orientation.

• 5) Preferential stabilization of the transition state: Enzyme binds the transition state more tightly than the substrate or product.

• 6) Protein conformational changes: Structural changes in the enzyme can facilitate catalysis.

Reaction Coordinate Diagram:

The reaction coordinate diagram illustrates the reduction in activation energy ($\Delta G^{\ddagger}$) for the catalyzed reaction compared to the uncatalyzed reaction.

$E + S \xrightleftharpoons[k_{-1}]{k_1} ES \xrightleftharpoons[k_{-2}]{k_2} EP \xrightleftharpoons[k_{-3}]{k_3} E + P$

Coenzymes, Vitamins, and Essential Metals

Coenzyme or Cofactor Function in Catalysis

Many enzymes require non-protein components for catalytic activity:

• Coenzymes: Organic molecules (often derived from vitamins) that assist in enzyme-catalyzed reactions.

• Cofactors: Inorganic ions or metal ions required for enzyme activity.

Vitamin	Coenzyme	Reaction involving the coenzyme
Thiamine (B1)	Thiamine pyrophosphate	Activation and transfer of aldehydes
Riboflavin (B2)	Flavin mononucleotide, flavin adenine dinucleotide	Oxidation-reduction
Niacin (B3)	Nicotinamide adenine dinucleotide, nicotinamide adenine dinucleotide phosphate	Oxidation-reduction
Pantothenic acid (B5)	Coenzyme A	Acyl group activation and transfer
Pyridoxine (B6)	Pyridoxal phosphate	Various reactions involving amino acid activation
Biotin	Biotin	CO2 activation and transfer
Lipoic acid	Lipoamide	Acyl group activation; oxidation-reduction
Folic acid	Tetrahydrofolate	Activation and transfer of single-carbon functional groups
Vitamin B12	Adenosyl cobalamin, methyl cobalamin	Isomerization and methyl group transfers

Metal Ions in Enzymes

Metal ions serve as essential cofactors in many enzymes, often participating directly in catalysis or substrate binding.

Metal	Example of Enzymes	Role of Metal
Fe	Cytochrome oxidase	Oxidation-reduction
Cu	Ascorbic acid oxidase	Oxidation-reduction
Zn	Alcohol dehydrogenase	Helps bind NAD+
Mn	Histidine ammonia lyase	Aids in catalysis by electron withdrawal
Co	Glutamate mutase	Co is part of cobalamin coenzyme
Ni	Urease	Catalytic site
Mo	Xanthine oxidase	Oxidation-reduction
V	Nitrate reductase	Oxidation-reduction
Se	Glutathione peroxidase	Replaces S in one cysteine in active site
Mg	Many kinases	Helps bind ATP

Enzyme Inhibition

Drugs, Toxins, and Enzymatic Activity

Enzyme inhibitors are important in pharmacology and toxicology. They can be classified as reversible or irreversible inhibitors based on their mode of interaction with the enzyme.

• Reversible inhibitors: Bind noncovalently and can dissociate from the enzyme.

• Irreversible inhibitors: Bind covalently, permanently inactivating the enzyme.

Types of Reversible Inhibition

• Competitive inhibition: Inhibitor competes with substrate for binding at the active site. Substrate can be processed, but not the inhibitor.

• Uncompetitive inhibition: Inhibitor binds only to the enzyme-substrate (ES) complex, at a site distinct from the active site, reducing catalytic activity.

• Mixed (noncompetitive) inhibition: Inhibitor can bind to either the free enzyme or the ES complex, usually at a site different from the substrate binding site, affecting both substrate binding and catalysis.

Irreversible Inhibition

Irreversible inhibitors form covalent bonds with enzymes, often at the active site, leading to permanent loss of activity.

• Example: Diisopropyl fluorophosphate (DFP) binds to the active site serine of acetylcholinesterase, blocking nerve conduction and causing paralysis.

The Regulation of Enzyme Activity

Controlling Enzyme Functions in the Cellular Context

Cells regulate enzyme activity through several mechanisms to maintain metabolic balance and respond to environmental changes.

• Substrate level control: Reaction rate increases with substrate concentration, but large changes are needed for significant regulation.

• Regulation at committed steps: Feedback inhibition or activation at key control points, often mediated by allosteric enzymes, is an efficient means to maintain homeostasis.

• Covalent modification: Enzyme activity can be regulated by reversible (e.g., phosphorylation) or irreversible (e.g., zymogen activation) covalent changes.

Feedback Inhibition

In metabolic pathways, the end product can inhibit an enzyme at an early step, preventing overproduction and conserving resources.

Allostery

• Allosteric enzymes: Usually multisubunit proteins that change conformation upon binding substrates or effector molecules.

• Homoallostery: Cooperativity in substrate binding (e.g., hemoglobin).

• Heteroallostery: Regulation by nonsubstrate effector molecules (activators or inhibitors).

Covalent Modifications Used to Regulate Enzyme Activity

Reversible and Irreversible Modifications

• Phosphorylation: Addition of phosphate groups by protein kinases; reversible by dephosphorylation via protein phosphatases. Acts as an on/off switch for enzyme activity.

• Zymogen activation: Irreversible activation of enzymes by proteolytic cleavage (e.g., digestive enzymes, blood clotting factors).

Nonprotein Biocatalysts: Catalytic Nucleic Acids

Ribonucleic Acids as Enzymes

Some RNA molecules, called ribozymes, can catalyze chemical reactions. This discovery supports the hypothesis that early life forms may have relied on RNA for both genetic information and catalytic activity (the "RNA World" hypothesis).

• Ribozymes: Catalytic RNA molecules capable of self-replication and catalysis.

• Significance: Suggests that RNA played a central role in early evolution before the emergence of DNA and protein enzymes.