Enzymes and biocatalysis

Last updated: January 3, 2023

Summarytoggle arrow icon

Enzymes are proteins that act on substrates, catalyzing chemical reactions within the cell. Enzymes are specific in the sense that each enzyme only reacts with a few closely related substrates. Some enzymes require cofactors (biotin, lipoamide, cobalamin) to function properly. Enzymes can become denatured by changes in temperature or pH. Enzymes are classified as oxidoreductases, transferases, hydrolases, lyases, ligases, and isomerases, based on the type of reaction they catalyze. Enzyme kinetics is the study of enzyme reaction rates, which are determined using the Michaelis-Menten and Lineweaver-Burk equations. These equations can also be used to evaluate how different types of enzyme inhibitors affect the reaction rate. Enzyme deficiencies can result in severe diseases such as Lesch-Nyhan syndrome, Gaucher disease, and phenylketonuria.

Overviewtoggle arrow icon

  • Complex proteins that catalyze chemical reactions
  • Act on substrates that can either be cleaved or joined to form a new product (e.g., carbonic anhydrase enzyme → CO2 + H20 ⇄ H2CO3)
  • Essential for life; if enzymes did not exist, cellular reactions would not occur fast enough to sustain life.
  • Enzyme deficiencies can result in severe diseases (e.g., Lesch-Nyhan syndrome).
  • Enzyme name is usually based on the reaction catalyzed plus the suffix “-ase”: e.g., for the enzyme that adds hydroxyl groups (OH-) is formed as follows: hydroxyl + -ase hydroxylase

General characteristics of enzymestoggle arrow icon

General characteristics

  • Active site: binding site for a specific substrate on a specific enzyme
  • Specificity
    • Enzymes are highly specific for their substrate and product.
    • Some exceptions include proteases that break down proteins to peptides in the digestive system.
  • Rate: enzymes catalyze reactions by a factor of 106–1011
  • Coenzymes
    • Many enzymes require coenzymes (e.g., biotin) that allow them to perform their action on a substrate.
    • Usually small organic molecules derived from metal ions or vitamins
  • Thermodynamics
    • Enzymes do not affect the energy level of substrates or products (free energy released remains the same).
    • Enzymes are able to decrease the energy of activation required to start a reaction.
    • The velocity of enzymatic reactions increases with temperature (up to 37o C in humans).
  • pH
    • Each enzyme has a specific pH at which it can achieve maximum velocity (Vmax).
    • Alterations in pH can cause denaturation of enzymes (specific to each enzyme).
    • Example: Pepsin works best in acidic environment like the stomach (pH ∼1.5–2) and it is inactivated in the duodenum when bicarbonate is released from the pancreas, increasing the pH to > 7.

Gibbs energy

Energy (∆G) for enzymatic reactions usually comes from the break down of ATP or GTP bonds (hydrolysis). Enzymatic reactions can occur spontaneously or nonspontaneously. The following are relationships between energy and enzymatic activity.

  • Exergonic: Energy (∆G) < 0: Reactions can occur spontaneously (often irreversible).
  • Endergonic: Energy (∆G) > 0: Reactions require energy to occur (from ATP or GDP).
  • Balanced reaction: Energy (∆G) = 0: The reaction is at equilibrium (reversible).

Classes of enzymestoggle arrow icon

Overview of enzyme classes
Enzyme class Function Subclass Examples
  • Dehydrogenases: catalyze oxidation-reduction reactions
  • Oxidases
  • Oxygenases
  • Hydroxylases: transfer hydroxyl groups (OH) onto substrates
  • Transfer functional groups
  • Kinases: transfer phosphate groups from a high energy molecule (e.g., ATP, ADP) onto substrates
  • Phosphorylases
    • Add inorganic phosphate to substrates
    • Do not require any energy source (e.g., ATP)
  • Glycosyltransferases
  • Cleave covalent bonds by adding water
  • Phosphatases: remove phosphate groups from substrates
  • Peptidases
  • Nucleosidases
  • Esterases
  • Aldolases
  • Decarboxylases
  • Dehydratases
  • Converts a substrate into its isomer
  • Mutases: move functional groups within a molecule
  • Epimerases
Ligases (sometimes called synthetases) [1]
  • Carboxylases
    • Transfer carbon dioxide groups (CO2)
    • Require biotin

Energy carrierstoggle arrow icon

Overview of energy carriers
Base molecule Transferred group Carrier of energy Released energy Metabolic site Molecular structure


  • -31 KJ/mol
  • Ubiquitous energy source


  • -31 KJ/mol


  • PKr
  • -43 KJ/mol


  • Thioester
  • -36 KJ/mol


  • PEP
  • -62 KJ/mol

Cofactorstoggle arrow icon

For more information, see “Vitamins.”

Overview of cofactors
Cofactor Vitamin Structure Reaction
Thiamine pyrophosphate (TPP)
  • Electron transfer
  • Electron transfer
  • Electron transfer
  • Electron transfer
Coenzyme A
  • Acyl group transfer
Pyridoxal phosphate
  • Methyl group transfer
  • Alkyl group transfer
S-Adenosylmethionine (SAM)
  • N/A
  • Methyl group transfer
  • N/A
Ascorbic acid
  • N/A
  • Electron transfer
  • Oxygen atom transfer
  • N/A

Rate-limiting enzymestoggle arrow icon

Overview of rate-limiting enzymes
Pathway Enzyme Stimulation Inhibition
  • Fructose-2,6-biphosphate
  • AMP
  • Fructose-1,6-biphosphatase
  • Fructose-2,6-biphosphate
  • AMP

Citric acid cycle



Pentose phosphate pathway (HMP shunt)

De novo pyrimidine synthesis
  • UTP
De novo purine synthesis
Urea cycle
  • N/A
Fatty acid synthesis


  • Carnitine acyltransferase I
  • N/A
  • N/A
Cholesterol synthesis

Enzyme kineticstoggle arrow icon

Michaelis-Menten kinetics

Description of an enzymatic reaction

  • Enzymatic reactions with a hyperbolic curve (most common, e.g., alcohol dehydrogenase in ethanol oxidation): E + S ⇄ ES → E + P
    • [E] = enzyme
    • [S] = substrate
    • [P] = product
    • [V] = velocity
  • A sigmoid curve indicates cooperativity (e.g., oxygen binding to hemoglobin)

Enzymatic reactions with a sigmoidal kinetic are indicative of cooperative binding (e.g., oxygen to hemoglobin).

Michaelis-Menten equation

  • Equation: v = Vmax [S] / (Km + [S])
  • Maximum velocity (Vmax)
    • Maximum rate at which an enzyme can catalyze a reaction
    • Directly proportional to the enzyme concentration
      • The only way to Vmax is to increase [E] (cells achieve this, e.g., by increasing gene expression of a given enzyme)
      • ↑ Enzyme concentration → Vmax
      • Noncompetitive inhibitors → ↓ [E] → Vmax
  • Michaelis constant: (Km): the substrate concentration at which half of the active sites of the enzymes are bound to the substrate
    • Reaction velocity is ½ of Vmax when the Michaelis constant concentration is reached
    • Inversely proportional to the affinity of the enzyme for the substrate: ↑ enzyme affinity → ↓ Km
  • Michaelis-Menten plot: a way of modeling Michaelis-Menten enzyme kinetics that relates the concentration of the substrate to reaction rate

Lineweaver-Burk equation and plot


  • The Lineweaver-Burk equation is a double reciprocal of the Michaelis-Menten equation, where V = Vmax [S] / Km + [S] (if [E] remains constant), becomes 1 / v = Km / Vmax× 1/[S] + 1 / Vmax.
  • Represents enzyme kinetics in a linear graph rather than a hyperbola
  • Equation is particularly important to determine the effect of drugs on enzymes


  • Intercept with y-axis: : 1/Vmax, the further from zero, the lower Vmax
  • Intercept with x-axis: : 1/-Km : the closer to zero, the lower the affinity and the higher the Km
  • Slope: Km/Vmax

Very efficient and kompetent: On the Lineweaver-Burk plot, Vmax usually represents the efficacy of a drug on the y-axis and Km represents the potency on the x-axis.

Drug-response dynamics

For details, see “Pharmacodynamics.”

Overview of drug-response dynamics
Parameter Uncompetitive inhibitors

Noncompetitive inhibitors

Competitive inhibitors (reversible)

Competitive inhibitors (irreversible)

Similar to the substrate
  • No
  • No
  • Yes
  • Yes
Effect of increased [S]
  • None
  • None
  • Inhibition can be overcome
  • None
Binding site
  • Binds to enzyme-substrate complex
  • Do not bind to active site
  • Binds to both the enzyme and enzyme-substrate complex
  • Reversible or irreversible
Effect on Km
  • Decreased
  • None
  • Increased
  • None
Effect on Vmax
  • Decreased
  • Decreased
  • None
  • Decreased
Pharmacodynamic effect

Uncompetitive inhibitors are enzyme inhibitors that bind to the enzyme-substrate complex, decreasing Km and Vmax.

Kompetitive Inhibition: Km Increases and Vmax remains unchanged. Nonkompetitive Inhibition: No Km change and Vmax decreases.

Only Companions meet on the waY: Competitive inhibitors meet on the Y-axis (same Vmax), noncompetitive do not.

Referencestoggle arrow icon

  1. Nomenclature Committee of IUB, Joint Commission on Biochemical Nomenclature. Nomenclature Committee of IUB (NC-IUB) IUB-IUPAC Joint Commission on Biochemical Nomenclature (JCBN) Newsletter 1984. Biosci Rep. 1984; 4 (2): p.177-180.doi: 10.1007/bf01120315 . | Open in Read by QxMD

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 Evidence-based content, created and peer-reviewed by physicians. Read the disclaimer