Enzyme inhibition refers to the ability to reduce or lose the activity of the enzyme, but does not cause the denaturation of the enzyme protein. Enzyme inhibition is mainly caused by changes in the chemical properties of the essential groups of the enzyme. Compounds that cause enzyme inhibition are called inhibitors. It should be noted that enzyme inhibition is different from enzyme inactivation, and inhibitors are also different from denaturants. Enzyme inhibition includes reversible inhibition and irreversible inhibition.

Reversible Inhibition

Reversible inhibition refers to the temporary loss of enzyme activity caused by the binding of inhibitors to enzyme proteins in a non-covalent manner. Reversible inhibitors can be removed by physical methods such as dialysis and can partially or completely restore enzyme activity. Reversible inhibition includes competitive inhibition, uncompetitive inhibition, non-competitive inhibition, and mixed inhibition.

• Competitive inhibition

The chemical structure of competitive inhibitors is similar to that of substrates, so they can compete with substrates and bind to enzyme active sites. When the competitive inhibitor binds to the active center of the enzyme, the substrate is excluded from the reaction center, resulting in the inhibition of the enzymatic reaction. Competitive inhibition can usually be eliminated by increasing the concentration of the substrate, that is, improving the competitiveness of the substrate.

Figure 1. Diagram of competitive inhibition.

• Non-competitive inhibition

Enzymes can bind to both substrates and non-competitive inhibitors, that is, there is no competition between substrates and non-competitive inhibitors. After the enzyme binds to the non-competitive inhibitor, it can also bind to the substrate, and after the enzyme binds to the substrate, it can also bind to the non-competitive inhibitor. However, in this case, the intermediate products of the enzymatic reaction cannot be further decomposed into products, so the enzyme activity is reduced.

Figure 2. Diagram of non-competitive inhibition.

Competitive inhibition can be overcome by sufficiently high concentrations of substrate, i.e., by out-competing the inhibitor. However, the apparent Km will increase as it takes a higher concentration of the substrate to reach the Km point, or half the Vmax. In non-competitive inhibition, Vmax will decrease due to the inability for the reaction to proceed as efficiently, but Km will remain the same as the actual binding of the substrate, by definition, will still function properly.

Figure 3. Michaelis–Menten saturation curve for an enzyme reaction showing the relation between the substrate concentration and reaction rate without inhibitor, with competitive inhibitor and non-competitive inhibitor.

• Uncompetitive inhibition

In uncompetitive inhibition, the inhibitor binds only to the substrate-enzyme complex. This type of inhibition causes Vmax to decrease and Km to decrease.

Figure 4. Diagram of uncompetitive inhibition.

• Mixed inhibition

In mixed inhibition, the inhibitor can bind to the enzyme at the same time as the enzyme's substrate. However, the binding of the inhibitor affects the binding of the substrate, and vice versa. This type of inhibition can be reduced, but not overcome by increasing concentrations of substrate.

Figure 5. Diagram of mixed inhibition.

Irreversible Inhibition

Irreversible inhibition means that the inhibitor covalently binds to the functional group of the active center of the enzyme, thus inhibiting the activity of the enzyme. Irreversible inhibitors cannot be removed by physical methods such as dialysis and restore enzyme activity.

Irreversible inhibitors often contain reactive functional groups such as nitrogen mustards, aldehydes, haloalkanes, alkenes, Michael acceptors, phenyl sulfonates, or fluorophosphonates. These electrophilic groups react with amino acid side chains to form covalent adducts. Irreversible inhibition is different from irreversible enzyme inactivation. Irreversible inhibitors are generally specific for one class of enzyme and do not inactivate all proteins; they do not function by destroying protein structure but by specifically altering the active site of their target. Irreversible inhibitors display time-dependent inhibition and their potency therefore cannot be characterised by an IC50 value. This is because the amount of active enzyme at a given concentration of irreversible inhibitor will be different depending on how long the inhibitor is pre-incubated with the enzyme.

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