When an inhibitor binds to the active site this type of inhibitor is called?

Enzymes are required for most, if not all, processes required for life. Enzymes catalyse a reaction by reducing the activation energy needed for the reaction to occur. However, enzymes need to be tightly regulated to ensure that levels of the product do not rise to undesired levels. This is accomplished by enzyme inhibition.

Types of Inhibition

Reversible and irreversible inhibitors are chemicals which bind to an enzyme to suppress its activity. One method to accomplish this is to almost permanently bind to an enzyme. These types of inhibitors are called irreversible. However, other chemicals can transiently bind to an enzyme. These are called reversible. Reversible inhibitors either bind to an active site (competitive inhibitors), or to another site on the enzyme (non-competitive inhibitors).

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Competitive Inhibitors

Competitive inhibitors compete with the substrate at the active site, and therefore increase Km (the Michaelis-Menten constant). However, Vmax is unchanged because, with enough substrate concentration, the reaction can still complete. The graph plot of enzyme activity against substrate concentration would be shifted to the right due to the increase of the Km, whilst the Lineweaver-Burke plot would be steeper when compared with no inhibitor.

Non-Competitive Inhibitors

Non-competitive inhibitors bind to another location on the enzyme and as such decrease VMAX. However, KM is unchanged. This is demonstrated by a lower maximum on a graph plotting enzyme activity against substrate concentration and a higher y-intercept on a Lineweaver-Burke plot when compared with no inhibitor.

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Allosteric Inhibition

Allosteric enzymes display a sigmoidal curve in contrast to the hyperbolic curve displayed by Michaelis-Menten Enzymes. This is because most allosteric enzymes contain multiple sub-units which can affect each other when the substrate binds to the enzyme. Inhibition can affect either K0.5, which is the substrate concentration for half-saturation, Vmax or both. This results in a shift of the curve to the right, and in the case of reducing Vmax, shifts the curve down.

Allosteric enzymes have two states: a low affinity state dubbed the “T” state and the high affinity “R” state. Inhibitors work by preferentially binding to the T state of an allosteric enzyme, causing the enzyme to maintain this low affinity state.

This is extremely useful to limit the amount of an enzyme’s product, as the product can then go on to inhibit the same type of enzyme to ensure the amount of product is not excessive. This is known as feedback inhibition. For example, ATP allosterically inhibits pyruvate kinase to prevent increased formation of pyruvate, so less ATP is eventually formed. Additionally, phosphofructokinase is allosterically inhibited by citrate, an intermediate of the Kreb’s cycle. This means that glycolysis will be limited when there is high ATP generation from the Kreb’s cycle.

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Phosphorylation

Phosphorylation provides another mechanism by which enzymes can be inhibited. This typically occurs through the action of kinase enzymes, which can either inhibit or activate an enzyme depending on the situation. The kinase enzymes cleave off a phosphate group from ATP and binds it to the enzyme. In situations where this results in an increase in enzyme activity it creates a cascade reaction, allowing a large response to be generated from a small stimulus.

Zymogens

Enzymes can also be secreted in an inactive state, known as zymogens. Zymogens are a useful mechanism allowing enzymes to be safely transported to different locations, without the enzyme becoming active and performing its function along the way. They remain inactive due to an addition of amino acids in the protein. Therefore, to activate a zymogen, another enzyme must cleave off these additional amino acids. For example, chymotrypsinogen is synthesised by the pancreas, yet is inactivated and cannot function. Upon travelling to the intestines, another enzyme (trypsin) cleaves off the additional amino acids to produce the activated form, chymotripsin.

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[start-clinical]

Clinical Relevance - Pharmacokinetics

The most important clinical use of enzyme inhibition is the use of pharmaceutical drugs. ACE inhibitors are a common treatment for hypertension. Angiotensin I is converted to angiotensin II by the action of angiotensin converting enzyme (ACE). However, angiotensin II causes numerous effects resulting in increased blood pressure. Therefore, ACE inhibitors are designed to competitively inhibit the action of ACE, which results in less angiotensin II formation and lower blood pressure.

Alternatively, Penicillin irreversibly binds to the active site of an enzyme called DD-transpeptidase. DD-transpeptidase is responsible for the final step of bacterial cell wall synthesis. By inhibiting this enzyme, the bacteria can not synthesise a cell wall and therefore can not sustain life.

Cyanide is an example of a non-competitive inhibitor. Cyanide binds to the final enzyme in the electron transport chain, and prevents this enzyme from catalysing the reaction from oxygen to water. This prevents the flow of electrons down the electron transport chain and no ATP can be generated, which results in death.

[end-clinical]

Previously, we noted that enzymes are inactivated at high temperatures and by changes in pH. These are nonspecific factors that would inactivate any enzyme. The activity of enzymes can also be regulated by more specific inhibitors. Many compounds are poisons because they bind covalently to particular enzymes or kinds of enzymes and inactivate them (Table \(\PageIndex{1}\)).

An irreversible inhibitor inactivates an enzyme by bonding covalently to a particular group at the active site. The inhibitor-enzyme bond is so strong that the inhibition cannot be reversed by the addition of excess substrate. The nerve gases, especially Diisopropyl fluorophosphate (DIFP), irreversibly inhibit biological systems by forming an enzyme-inhibitor complex with a specific OH group of serine situated at the active sites of certain enzymes. The peptidases trypsin and chymotrypsin contain serine groups at the active site and are inhibited by DIFP.

A reversible inhibitor inactivates an enzyme through noncovalent, more easily reversed, interactions. Unlike an irreversible inhibitor, a reversible inhibitor can dissociate from the enzyme. Reversible inhibitors include competitive inhibitors and noncompetitive inhibitors. (There are additional types of reversible inhibitors.) A competitive inhibitor is any compound that bears a structural resemblance to a particular substrate and thus competes with that substrate for binding at the active site of an enzyme. The inhibitor is not acted on by the enzyme but does prevent the substrate from approaching the active site.

The degree to which a competitive inhibitor interferes with an enzyme’s activity depends on the relative concentrations of the substrate and the inhibitor. If the inhibitor is present in relatively large quantities, it will initially block most of the active sites. But because the binding is reversible, some substrate molecules will eventually bind to the active site and be converted to product. Increasing the substrate concentration promotes displacement of the inhibitor from the active site. Competitive inhibition can be completely reversed by adding substrate so that it reaches a much higher concentration than that of the inhibitor.

Studies of competitive inhibition have provided helpful information about certain enzyme-substrate complexes and the interactions of specific groups at the active sites. As a result, pharmaceutical companies have synthesized drugs that competitively inhibit metabolic processes in bacteria and certain cancer cells. Many drugs are competitive inhibitors of specific enzymes.

A classic example of competitive inhibition is the effect of malonate on the enzyme activity of succinate dehydrogenase (Figure \(\PageIndex{1}\)). Malonate and succinate are the anions of dicarboxylic acids and contain three and four carbon atoms, respectively. The malonate molecule binds to the active site because the spacing of its carboxyl groups is not greatly different from that of succinate. However, no catalytic reaction occurs because malonate does not have a CH2CH2 group to convert to CH=CH. This reaction will also be discussed in connection with the Krebs cycle and energy production.

Chemotherapy is the strategic use of chemicals (that is, drugs) to destroy infectious microorganisms or cancer cells without causing excessive damage to the other, healthy cells of the host. From bacteria to humans, the metabolic pathways of all living organisms are quite similar, so the search for safe and effective chemotherapeutic agents is a formidable task. Many well-established chemotherapeutic drugs function by inhibiting a critical enzyme in the cells of the invading organism.

An antibiotic is a compound that kills bacteria; it may come from a natural source such as molds or be synthesized with a structure analogous to a naturally occurring antibacterial compound. Antibiotics constitute no well-defined class of chemically related substances, but many of them work by effectively inhibiting a variety of enzymes essential to bacterial growth.

Penicillin, one of the most widely used antibiotics in the world, was fortuitously discovered by Alexander Fleming in 1928, when he noticed antibacterial properties in a mold growing on a bacterial culture plate. In 1938, Ernst Chain and Howard Florey began an intensive effort to isolate penicillin from the mold and study its properties. The large quantities of penicillin needed for this research became available through development of a corn-based nutrient medium that the mold loved and through the discovery of a higher-yielding strain of mold at a United States Department of Agriculture research center near Peoria, Illinois. Even so, it was not until 1944 that large quantities of penicillin were being produced and made available for the treatment of bacterial infections.

Penicillin functions by interfering with the synthesis of cell walls of reproducing bacteria. It does so by inhibiting an enzyme—transpeptidase—that catalyzes the last step in bacterial cell-wall biosynthesis. The defective walls cause bacterial cells to burst. Human cells are not affected because they have cell membranes, not cell walls.

Several naturally occurring penicillins have been isolated. They are distinguished by different R groups connected to a common structure: a four-member cyclic amide (called a lactam ring) fused to a five-member ring. The addition of appropriate organic compounds to the culture medium leads to the production of the different kinds of penicillin.

The penicillins are effective against gram-positive bacteria (bacteria capable of being stained by Gram’s stain) and a few gram-negative bacteria (including the intestinal bacterium Escherichia coli). They are effective in the treatment of diphtheria, gonorrhea, pneumonia, syphilis, many pus infections, and certain types of boils. Penicillin G was the earliest penicillin to be used on a wide scale. However, it cannot be administered orally because it is quite unstable; the acidic pH of the stomach converts it to an inactive derivative. The major oral penicillins—penicillin V, ampicillin, and amoxicillin—on the other hand, are acid stable.

Some strains of bacteria become resistant to penicillin through a mutation that allows them to synthesize an enzyme—penicillinase—that breaks the antibiotic down (by cleavage of the amide linkage in the lactam ring). To combat these strains, scientists have synthesized penicillin analogs (such as methicillin) that are not inactivated by penicillinase.

Some people (perhaps 5% of the population) are allergic to penicillin and therefore must be treated with other antibiotics. Their allergic reaction can be so severe that a fatal coma may occur if penicillin is inadvertently administered to them. Fortunately, several other antibiotics have been discovered. Most, including aureomycin and streptomycin, are the products of microbial synthesis. Others, such as the semisynthetic penicillins and tetracyclines, are made by chemical modifications of antibiotics; and some, like chloramphenicol, are manufactured entirely by chemical synthesis. They are as effective as penicillin in destroying infectious microorganisms. Many of these antibiotics exert their effects by blocking protein synthesis in microorganisms.

Initially, antibiotics were considered miracle drugs, substantially reducing the number of deaths from blood poisoning, pneumonia, and other infectious diseases. Some seven decades ago, a person with a major infection almost always died. Today, such deaths are rare. Seven decades ago, pneumonia was a dreaded killer of people of all ages. Today, it kills only the very old or those ill from other causes. Antibiotics have indeed worked miracles in our time, but even miracle drugs have limitations. Not long after the drugs were first used, disease organisms began to develop strains resistant to them. In a race to stay ahead of resistant bacterial strains, scientists continue to seek new antibiotics. The penicillins have now been partially displaced by related compounds, such as the cephalosporins and vancomycin. Unfortunately, some strains of bacteria have already shown resistance to these antibiotics.

Some reversible inhibitors are noncompetitive. A noncompetitive inhibitor can combine with either the free enzyme or the enzyme-substrate complex because its binding site on the enzyme is distinct from the active site. Binding of this kind of inhibitor alters the three-dimensional conformation of the enzyme, changing the configuration of the active site with one of two results. Either the enzyme-substrate complex does not form at its normal rate, or, once formed, it does not yield products at the normal rate. Because the inhibitor does not structurally resemble the substrate, the addition of excess substrate does not reverse the inhibitory effect.

Feedback inhibition is a normal biochemical process that makes use of noncompetitive inhibitors to control some enzymatic activity. In this process, the final product inhibits the enzyme that catalyzes the first step in a series of reactions. Feedback inhibition is used to regulate the synthesis of many amino acids. For example, bacteria synthesize isoleucine from threonine in a series of five enzyme-catalyzed steps. As the concentration of isoleucine increases, some of it binds as a noncompetitive inhibitor to the first enzyme of the series (threonine deaminase), thus bringing about a decrease in the amount of isoleucine being formed (Figure \(\PageIndex{2}\)).

An irreversible inhibitor inactivates an enzyme by bonding covalently to a particular group at the active site. A reversible inhibitor inactivates an enzyme through noncovalent, reversible interactions. A competitive inhibitor competes with the substrate for binding at the active site of the enzyme. A noncompetitive inhibitor binds at a site distinct from the active site.