Enzymes are protein-based molecules that operate as biocatalysts in the body by accelerating chemical reactions and processes. Enzymes are generally recognized to catalyze more than 5000 different types of biological reactions.
In the human body, enzymes catalyze all types of chemical reactions that are necessary for normal growth, blood clotting, the healing process, digestion, DNA replication, transcription, translation, and signal transduction.
All enzymes normally produce an enzyme-substrate complex by first binding with the substrate, which is subsequently transformed into the final product. The enzyme isremoved from the final product after the reaction, and it is now free to turn another identical substrate into the final product.
Many organic or synthetic substances attach to enzymes and alter their reaction rates. Enzyme activators and inhibitors are described as molecules that increase or decrease an enzyme’s activity, respectively.
Inhibitors change the catalytic action of the enzyme, causing it to slow down or, in some cases, stop catalysis. The research onthese inhibitors yields a wealth of knowledge regarding the mechanism and operation of enzymes.
The inhibition of enzyme activity can result in a number of modifications, including the correction of metabolic imbalance or the eradication of bacteria or pathogens. Because of this, enzyme inhibitors make up a large portion of therapeutic compounds, and biochemistry and pharmacology researchers have spent a lot of time discovering and improving these molecules.
The specificity and potency of an enzyme inhibitor are measured, and their high value ensures that the inhibitor has fewer side effects and is less harmful.
There are three different kinds of enzyme inhibitors-Reversible, irreversible, and allosteric. In addition to these common types of enzyme inhibition, allosteric inhibition andphosphorylation are other typeswhose mechanisms are completely distinct from those of the aforementioned inhibitions.
A. Reversible Inhibition
The name implies that the molecule that inhibits the enzyme is not covalently attached to the enzyme and, after some time, it separates from the enzyme, allowing us to obtainfree enzyme. The weak interactions that reversible enzyme inhibitors use to attach to the enzyme, such as hydrogen bonds, hydrophobic interactions, and ionic bonds, might break after a finite amount of time.
Dilution or dialyses are used to restore enzyme activity. The kind of binding with the enzyme and the enzyme-substrate complex affects the inhibition. Competitive, Non-Competitive, and Uncompetitive inhibitors are the three types ofreversible inhibitors.
Mechanism of reversible inhibition where E is enzyme, S is Substrate, EI is enzyme inhibitor complex, ES is Enzyme substrate complex, P is product, And I is Inhibitor
1. Competitive inhibition
Due to the inhibitor’s structural similarity to the substrate, it attaches to the active site in a competitive manner. However, when the concentration of the competitive inhibitor is greater than that of the substrate, it binds to the active binding site to form an enzyme inhibitor complex (EI), which ultimately prevents the formation of any product.
The idea that inhibitors lash away the substrate from the enzyme is completely confusing information. Instead, it is true that the inhibitor could bind to the enzyme-substrate complex and would force the substrate to separate from the enzyme using the thermodynamic principle, in which the concentration and affinity of the substrate/inhibitor and the enzyme govern the interaction between the two.
[NOTE: The maximal reaction velocity in a typical enzymatic reaction is known as Vmax, and the substrate concentration that is halfway to Vmax is known as Km (or the Michaelis-Menten constant). Km is an appropriate unit of measurement for determining the rate of reaction as substrate concentration increases. Higher substrate affinity is associated with lower Km values, and vice versa.]
Due to their competition for the same binding site, the substrate’s Km appears to increase when the inhibitor is present. The inhibition can be restored by exposing the competing inhibitor to a high concentration of substrate. The reaction’s Vmax remains constant, but the dissociation constant, kd, appears to have increased. The enzyme active site can also be located using competitive inhibitors.
Statins like atrovastatin, which block the HMG CoA reductase enzyme and are used to treat atherosclerosis as an anti-hyperlipidemic medication, are among common helpful pharmaceuticals that exhibit this type of inhibition. Gout is treated with the Xanthine Oxidase Inhibitor Allopurinol.
Combining methotrexate and sulfonamides inhibits the Dihydrofolate reductase enzyme. Angiotensin-converting enzyme inhibitors, such as captopril, enalapril, and ramipril, are prescribed to people with heart disease and high blood pressure.
2. Non-competitive inhibition
There is no structural similarity between the inhibitor and the substrate in non-competitive inhibition. The inhibitors bind to the enzyme at different places than the substrate binding site, resulting in the formation of enzyme-inhibitor (EI) and enzyme-inhibitor-substrate (EIS) complexes. Since the inhibitor and enzyme establish non-covalent bonds, the inhibition of the enzyme can be eliminated by just removing the inhibitor.
Simple non-competitive inhibition occurs when the affinities of the enzyme and enzyme inhibitor complexes for the substrate are identical, but the product produced by the enzyme-inhibitor-substrate complex is very low. When the apparent affinity of the enzyme for substrate is affected by inhibitor binding, non-competitive inhibition becomes more complex.
The inhibitor slows down the reaction by attaching to a different spot on the same enzyme and altering its overall shape so that the substrate can fit into it as before. The response is delayed but never completely stopped.
Instead of reducing the amount of substrate bound to the enzyme, non-competitive inhibition reduces the rate of enzyme turnover. Specific non-competitive enzyme inhibition is a term used by some authors to describe this type of inhibition.
The behavior of a non-competitive inhibitor mimics the removal of the active enzyme from the solution, lowering Vmax. The double reciprocal plotshows this graphically, with Km remaining constant but Vmax changes.
Alcohol and narcotic medicines are two very typical examples of substances that non-competitively block acid phosphate. Pepstatin is one example of a powerful inhibitor of aspartyl proteases.
3. Uncompetitive Inhibitors
Uncompetitive inhibitors bind to the enzyme-substrate complex exclusively. It doesn’t bind to the unbound enzyme. Because of the structural deformation of the active site, they are able to prevent biological agonist molecules from binding to the active site. Pharmacological action is not produced if the agonist molecule does not correctly attach to the active site. This type of inhibition caused a drop in both Vmax and Km.
Contrary to the situation for competitive inhibition, adding substrate does not undo the effects of an uncompetitive inhibitor since its binding does not conflict with that of the uncompetitive inhibitor.
The inhibitor must interfere with the enzyme’s catalytic activity but not its substrate binding in order to cause uncompetitive inhibition. To imagine this for single-substrate enzymes is challenging. Uncompetitive inhibition is actually only important for multisubstrate enzymes.
4. Mixed Inhibition
An inhibitor may bind to both the enzyme and the enzyme-substrate complex, resulting in mixed inhibition. The affinities with which the inhibitor binds to the two different states, however, may differ. When the enzyme is free compared to the enzyme-substrate complex, it may bind more strongly, or the opposite may be true.
Both competitive and non-competitive inhibitions are present in the inhibition. Mixed inhibition will turn into non-competitive inhibition if the inhibitor has an identical affinity for the enzyme in both its free and substrate-bound forms. In mixed inhibition, the substrate-binding active site of the enzyme is not the place where the inhibitor binds.
A mixed inhibitor, presumably, binds to enzyme sites involved in both substrate binding and catalysis. Examples of mixed inhibitors include metal ions, which unlike competitive inhibitors do not directly compete with substrates for binding to an enzyme active site.
The apparent values of Km and Vmax are changed by the presence of the inhibitor, just like in non-competitive inhibition.
B. Irreversible inhibition
Irreversible inhibitors attach to enzymes and chemically modify them by forming covalent bonds with them. Key amino acids that are necessary for activity are present in the structure of enzymes, and irreversible inhibitors firmly bind to these amino acid side chains. Inhibitors bind to enzymes covalently (firmly) and irreversibly.
So it is unable to separate from the enzyme. Dialysis or elevating concentration cannot restore an enzyme’s activity. They are also known as suicide inhibitors because they bind to enzymes for an extended period of time and do not readily detach from their binding sites, which can lead to toxicity and cause death.
Examples of the best irreversible inhibitors include: 1. Alcohol dehydrogenase inhibitor disulfiram. Alcoholism is treated with disulfiram. 2. Acetylcholine esterase inhibitor malathion is a component of organophosphorus insecticides.
Suicide Inhibition
Another form of irreversible inhibition is suicide inhibition. In this instance, the inhibitor substance is changed by the target enzyme into a reactive form in its active site. They are also referred to as transition state analogues or mechanism-based inhibitors.
For instance, the analogue of ornithine DFMO [-difluromethyl ornithine] inhibits ornithine decarboxylase. Coxygenase is inhibited by aspirin.
C. Allosteric inhibition
This sort of enzyme inhibition occurs when several enzymes function sequentially to catalyze processes along a pathway, and the last product may be to blame for preventing the first enzyme in the series from acting. The ultimate end product from the substrate molecule causes an entirely new structural change in the inhibitor.
An enzyme that causes such inhibition is known as an allosteric enzyme, and the inhibition itself is referred to as allosteric inhibition. The allosteric site, which is an additional site that some enzymes contain in addition to the active site, is a key target for research associates and medicinal chemists.
It comes in two types: positive allosteric, which results in an increase in enzyme activity, and negative allosteric, where enzyme activity is decreased.
The Michaelis-Menten model does not apply to allosteric enzymes. They have two or more active sites as well as more than two subunits. Instead of adhering to Michaelis-Menten kinetics, they follow sigmoidal kinetics.
Under intracellular conditions, allosteric enzymes often have an irreversible reaction. They typically have substantially higher molecular weights and more intricate configurations. Some of them are stable at body temperature or ambient temperature but unstable at zero degrees.
Allosteric regulated enzyme rates can be precisely controlled by subtle changes in substrate level. With modest doses of a negative allosteric effector, it is also feasible to “switch an enzyme off” by shifting the apparent Km to values significantly higher than the in vivo substrate level.
D. Phosphorylation
Phosphorylation is an essential enzyme inhibitor that can change the activity of proteins after they are produced. By means of particular enzymes called kinases, a protein receives a phosphate group. The phosphate group is often supplied by ATP, the cell’s energy transporter. An organic molecule is phosphorylated when a phosphoryl group (PO3-) is chemically added to it.
In the body, phosphorylation takes three different forms- oxidative phosphorylation, protein phosphorylation, and glucose phosphorylation. Glucose phosphorylation can be promoted by the binding of fructose-6-phosphate and decreased by the binding of fructose-1-phosphate.
Glycogen synthase is triggered by the effector glucose 6 phosphate’s allosteric activation, and glucose 6 phosphate may prevent cyclic AMP-driven protein kinase from phosphorylating glycogen synthase.
Both prokaryotic and eukaryotic organisms engage in the crucial regulatory mechanism known as reversible phosphorylation of proteins. Many enzymes and receptors undergo reversible phosphorylation, which alters their structural conformation and either activates or deactivates them. Activation of the tumor suppressor protein p53 can cause cell cycle arrest, which can be reversed in some cases.
The way a cell accumulates and releases chemical energy is through oxidative phosphorylation. The chemical reactions take place inside the mitochondria in eukaryotic cells.The electron transport chain and chemiosmosis processes make up oxidative phosphorylation.
References
- Voet, D., Voet, J., & Pratt, C. (2016).Fundamentals of Biochemistry. New York: Wiley.
- Devlin, T.Textbook of biochemistry. Hoboken, NJ: Wiley-Liss.
- Patadiya, Nikunj & Panchal, Nikita & Vaghela, Vipul. (2021). A REVIEW ON ENZYME INHIBITORS. International Research Journal of Pharmacy. 12. 12. 10.7897/2230-8407.1206145.
- Sahu, Supriya & Pathak, Kalyani & Gogoi, Urvashee & Das, Aparoop & Saikia, Riya. (2020). Enzyme Inhibition. 10.22271/ed.book.795.