Enzymes: Biochemistry's Catalysts

• Enzymes are studied in biochemistry and have been crucial to the field
• The study of enzymes dates back to the late 1700s
• Enzymes increase reaction rates without being used up
• Most enzymes are globular proteins
• Some RNA molecules (ribozymes and ribosomal RNA) can also catalyze reactions

Contributors to Enzyme Study

• Eduard Buchner demonstrated that enzymes can work even outside of cells
• James Sumner crystallized urease enzyme and determined that enzymes are proteins
• J. B. S. Haldane wrote about enzymes and suggested weak bonding interactions are important in catalysis

Cofactors and Coenzymes

• Most enzymes are proteins, except for a small group of catalytic RNA molecules

• Some require additional molecules called cofactors or coenzymes to be fully functional

• Apoenzyme: The inactive protein part of an enzyme

• Cofactor: A non-protein component that can be either an inorganic metal ion or an organic molecule (coenzyme).

• Holoenzyme: A complete, active enzyme, which includes the apoenzyme and cofactor.

Inorganic Metal Ions

• Some metal ions serve as cofactors for enzymes:
  • Copper (Cu2+) is a cofactor for cytochrome oxidase.
  • Magnesium (Mg2+) is a cofactor for hexokinase, glucose 6-phosphatase, and pyruvate kinase.
  • Iron (Fe2+ or Fe3+) is a cofactor for cytochrome oxidase, catalase, and peroxidase

Organic Molecules of Enzymes (Coenzymes)

• Biocytin is involved in the transfer of carbon dioxide molecules, and its dietary precursor is biotin
• Pyridoxal phosphate is involved in the transfer of amino groups, and its dietary precursor is vitamin B6
• Many vitamins serve as dietary precursors for coenzymes in the body

Enzyme Nomenclature

• IUBMB (International Union of Biochemistry and Molecular Biology) assigns systematic names and Enzyme Commission (EC) numbers

• Example: ATP + D-Glucose➔ ADP + D-Glucose-6-Phosphate and the enzyme catalyzing named ATP:Glucose phosphotransferase

  • it's EC number is 2.7.1.1

• Enzyme Commission (EC) number:

  • 1st digit: Enzyme class name.
  • 2nd digit: Subclass.
  • 3rd digit: Acceptor group category.
  • 4th digit: Acceptor molecule

Enzyme Classes

• Oxidoreductases: Transfer electrons (hydride ions or H atoms)
• Transferases: Transfer functional groups
• Hydrolases: Hydrolyze bonds by adding water
• Lyases: Cleave C-C, C-O, C-N, or other bonds by elimination, forming double bonds or rings
• Isomerases: Transfer groups within a molecule to yield isomeric forms
• Ligases: Form C-C, C-S, C-O, and C-N bonds through condensation, coupled with ATP cleavage
• Translocases: Facilitate movement of molecules or ions across membranes

How Enzymes Work

• Enzymes function by increasing reaction rates but do not affect reaction equilibria
• Reactions can be described using a reaction coordinate diagram, where free energy is plotted against the reaction progress

Reaction Coordinate Diagram

• Ground state: The starting point for either the forward or reverse reaction

• Chemists define a standard set of conditions (298 K, 1 atm, 1 M) to describe free-energy changes for reactions

• Biochemical standard free-energy change (∆G'°) is defined at pH 7.0

• A negative ∆G’° indicates that the reaction is exergonic, favoring product formation

• There is an energy barrier between reactants (S) and products (P)

• Transition state: The highest energy point where decay to either the S or P state is equally probable

• Activation energy (∆G‡) is the difference between the ground state and transition state energy levels

• Higher activation energy corresponds to a slower reaction

• Catalysts lower activation energies to enhance reaction rates

• Rate-limiting step: The step with the highest activation energy determines the overall reaction rate

Enzymes Speed Up Reactions By Reducing Activation Energy

• Reaction coordinate diagram comparing enzyme-catalyzed and uncatalyzed reactions shows that ES and EP intermediates occupy minima in the energy progress curve of the enzyme catalyzed reaction.
• AG‡ uncat and AG‡ cat correspond to the activation energy for uncatalyzed and catalyzed reactions
• Activation energy is lower when the enzyme catalyzes the reaction
• Enzymes bind transition states better than substrates for faster reactions

How to Lower Activation Energy

• Linus Pauling proposed in 1946 that enzyme active sites are complementary to the transition state

  • Enzymes bind transition states better than substrates because active sites are designed to fit transition states of reactions

• Stronger interactions lower activation barrier.

• In catalyzed reactions, the enzyme uses the binding energy of substrates to organize the reactants to a fairly rigid ES complex

• Entropy cost is paid during binding

• A rigid reactant complex to transition state conversion is entropically neutral

Enzyme Specificity

• Binding energy provides energy for catalysis and gives an enzyme its specificity, the ability to discriminate between a substrate and a competing molecule
• Enzymes are able to catalyze specific reactions and exclude others because a well designed enzyme active site will only have functional groups arranged optimally to form weak interactions with a particular substrate in the transition state

Biocatalysis vs Inorganic Catalysts

• Enzymes make the desired reaction most favorable.

• Biocatalysis has many advantages:

  • Greater reaction specificity (fewer side products)
  • Milder reaction conditions (pH ~ 7, 37°C)
  • Higher reaction rates
  • Capacity for regulation
  • Control of biological pathways

Catalytic Mechanisms

• Enzymes may use one or a combination of the following:

  • Acid-base catalysis
  • Covalent catalysis
  • Metal ion catalysis

Acid-Base Catalysis

• Many organic reactions that are used to model biochemical processes are promoted by proton donors (general acids) or proton acceptors (general bases)

• Some active sites of enzymes contain amino acid functional groups that act as proton donors or acceptors:

  • These groups are precisely positioned to allow for the transfer of protons
  • Enhances reactions rates by 102 to 105 fold
  • This type of catalysis occurs on the vast majority of enzymes

Covalent Catalysis

• Involves the formation of a transient covalent bond between the enzyme and substrate
• Requires a nucleophile on the enzyme such as serine, thiolate, amine, or carboxylate

Metal Ion Catalysis

• A metal ion is bound to the enzyme
• Stabilizes negative charges to facilitate the binding of a substrate