Anenzyme (/ˈɛnzaɪm/) is aprotein that acts as a biologicalcatalyst by acceleratingchemical reactions. Themolecules upon which enzymes may act are calledsubstrates, and the enzyme converts the substrates into different molecules known asproducts. Almost allmetabolic processes in thecell needenzyme catalysis in order to occur at rates fast enough to sustain life.[1]: 8.1 Metabolic pathways depend upon enzymes to catalyze individual steps. The study of enzymes is calledenzymology and the field ofpseudoenzyme analysis recognizes that during evolution, some enzymes have lost the ability to carry out biological catalysis, which is often reflected in theiramino acid sequences and unusual 'pseudocatalytic' properties.[2][3]
Enzymes are known to catalyze more than 5,000 biochemical reaction types.[4]
Other biocatalysts includecatalytic RNA molecules, also calledribozymes. They are sometimes described as atype of enzyme rather than beinglike an enzyme, but even in the decades since ribozymes' discovery in 1980–1982, the wordenzyme alone often means the protein type specifically (as is used in this article). A third category of biocatalysts is constituted by thosebiomolecular condensates that have catalytic ability.[5]
Like all catalysts, enzymes increase thereaction rate by lowering itsactivation energy. Some enzymes can make their conversion of substrate to product occur many millions of times faster. An extreme example isorotidine 5'-phosphate decarboxylase, which allows a reaction that would otherwise take millions of years to occur in milliseconds.[6][7] Chemically, enzymes are like any catalyst and are not consumed in chemical reactions, nor do they alter theequilibrium of a reaction. Enzymes differ from most other catalysts by being much more specific. Enzyme activity can be affected by other molecules:inhibitors are molecules that decrease enzyme activity, andactivators are molecules that increase activity. Many therapeuticdrugs andpoisons are enzyme inhibitors. An enzyme's activity decreases markedly outside its optimaltemperature andpH, and many enzymes are (permanently)denatured when exposed to excessive heat, losing their structure and catalytic properties.
Some enzymes are used commercially, for example, in the synthesis ofantibiotics. Some household products use enzymes to speed up chemical reactions: enzymes inbiological washing powders break down protein, starch orfat stains on clothes, and enzymes inmeat tenderizer break down proteins into smaller molecules, making the meat easier to chew.
Etymology and history
By the late 17th and early 18th centuries, the digestion ofmeat by stomach secretions[8] and the conversion ofstarch tosugars by plant extracts andsaliva were known but the mechanisms by which these occurred had not been identified.[9]
French chemistAnselme Payen was the first to discover an enzyme,diastase, in 1833.[10] A few decades later, when studying thefermentation of sugar toalcohol byyeast,Louis Pasteur concluded that this fermentation was caused by avital force contained within the yeast cells called "ferments", which were thought to function only within living organisms. He wrote that "alcoholic fermentation is an act correlated with the life and organization of the yeast cells, not with the death or putrefaction of the cells."[11]
In 1877, German physiologistWilhelm Kühne (1837–1900) first used the termenzyme, which comes from Ancient Greekἔνζυμον (énzymon)'leavened, in yeast', to describe this process.[12] The wordenzyme was used later to refer to nonliving substances such aspepsin, and the wordferment was used to refer to chemical activity produced by living organisms.[13]
Eduard Buchner
Eduard Buchner submitted his first paper on the study of yeast extracts in 1897. In a series of experiments at theUniversity of Berlin, he found that sugar was fermented by yeast extracts even when there were no living yeast cells in the mixture.[14] He named the enzyme that brought about the fermentation of sucrose "zymase".[15] In 1907, he received theNobel Prize in Chemistry for "his discovery of cell-free fermentation". Following Buchner's example, enzymes are usually named according to the reaction they carry out: the suffix-ase is combined with the name of thesubstrate (e.g.,lactase is the enzyme that cleaveslactose) or to the type of reaction (e.g.,DNA polymerase forms DNA polymers).[16]
The biochemical identity of enzymes was still unknown in the early 1900s. Many scientists observed that enzymatic activity was associated with proteins, but others (such as Nobel laureateRichard Willstätter) argued that proteins were merely carriers for the true enzymes and that proteinsper se were incapable of catalysis.[17] In 1926,James B. Sumner showed that the enzymeurease was a pure protein and crystallized it; he did likewise for the enzymecatalase in 1937. The conclusion that pure proteins can be enzymes was definitively demonstrated byJohn Howard Northrop andWendell Meredith Stanley, who worked on the digestive enzymespepsin (1930),trypsin andchymotrypsin. These three scientists were awarded the 1946 Nobel Prize in Chemistry.[18]
The discovery that enzymes could be crystallized eventually allowed their structures to be solved byx-ray crystallography. This was first done forlysozyme, an enzyme found in tears, saliva andegg whites that digests the coating of some bacteria; the structure was solved by a group led byDavid Chilton Phillips and published in 1965.[19] This high-resolution structure of lysozyme marked the beginning of the field ofstructural biology and the effort to understand how enzymes work at an atomic level of detail.[20]
Classification and nomenclature
Enzymes can be classified by two main criteria: eitheramino acid sequence similarity (and thus evolutionary relationship) or enzymatic activity.
Enzyme activity. An enzyme's name is often derived from its substrate or the chemical reaction it catalyzes, with the word ending in-ase.[1]: 8.1.3 Examples arelactase,alcohol dehydrogenase andDNA polymerase. Different enzymes that catalyze the same chemical reaction are calledisozymes.[1]: 10.3
TheInternational Union of Biochemistry and Molecular Biology have developed anomenclature for enzymes, theEC numbers (for "Enzyme Commission"). Each enzyme is described by "EC" followed by a sequence of four numbers which represent the hierarchy of enzymatic activity (from very general to very specific). That is, the first number broadly classifies the enzyme based on its mechanism while the other digits add more and more specificity.[21]
EC 7,Translocases: catalyze the movement of ions or molecules across membranes, or their separation within membranes.
These sections are subdivided by other features such as the substrate, products, andchemical mechanism. An enzyme is fully specified by four numerical designations. For example,hexokinase (EC 2.7.1.1) is a transferase (EC 2) that adds a phosphate group (EC 2.7) to a hexose sugar, a molecule containing an alcohol group (EC 2.7.1).[22]
Sequence similarity. EC categories donot reflect sequence similarity. For instance, two ligases of the same EC number that catalyze exactly the same reaction can have completely different sequences. Independent of their function, enzymes, like any other proteins, have been classified by their sequence similarity into numerous families. These families have been documented in dozens of different protein and protein family databases such asPfam.[23]
Non-homologous isofunctional enzymes. Unrelated enzymes that have the same enzymatic activity have been callednon-homologous isofunctional enzymes.[24]Horizontal gene transfer may spread these genes to unrelated species, especially bacteria where they can replace endogenous genes of the same function, leading to hon-homologous gene displacement.
Structure
Enzyme activity initially increases with temperature (Q10 coefficient) until the enzyme's structure unfolds (denaturation), leading to an optimalrate of reaction at an intermediate temperature.
Enzymes are generallyglobular proteins, acting alone or in largercomplexes. The sequence of the amino acids specifies the structure which in turn determines the catalytic activity of the enzyme.[25] Although structure determines function, a novel enzymatic activity cannot yet be predicted from structure alone.[26] Enzyme structures unfold (denature) when heated or exposed to chemical denaturants and this disruption to the structure typically causes a loss of activity.[27] Enzyme denaturation is normally linked to temperatures above a species' normal level; as a result, enzymes from bacteria living in volcanic environments such ashot springs are prized by industrial users for their ability to function at high temperatures, allowing enzyme-catalysed reactions to be operated at a very high rate.
Enzymes are usually much larger than their substrates. Sizes range from just 62 amino acid residues, for themonomer of4-oxalocrotonate tautomerase,[28] to over 2,500 residues in the animalfatty acid synthase.[29] Only a small portion of their structure (around 2–4 amino acids) is directly involved in catalysis: the catalytic site.[30] This catalytic site is located next to one or morebinding sites where residues orient the substrates. The catalytic site and binding site together compose the enzyme'sactive site. The remaining majority of the enzyme structure serves to maintain the precise orientation and dynamics of the active site.[31]
In some enzymes, no amino acids are directly involved in catalysis; instead, the enzyme contains sites to bind and orient catalyticcofactors.[31] Enzyme structures may also containallosteric sites where the binding of a small molecule causes aconformational change that increases or decreases activity.[32]
A small number ofRNA-based biological catalysts calledribozymes exist, which again can act alone or in complex with proteins. The most common of these is theribosome which is a complex of protein and catalytic RNA components.[1]: 2.2
Enzymes must bind their substrates before they can catalyse any chemical reaction. Enzymes are usually very specific as to whatsubstrates they bind and then the chemical reaction catalysed.Specificity is achieved by binding pockets with complementary shape, charge andhydrophilic/hydrophobic characteristics to the substrates. Enzymes can therefore distinguish between very similar substrate molecules to bechemoselective,regioselective andstereospecific.[33]
Some of the enzymes showing the highest specificity and accuracy are involved in the copying andexpression of thegenome. Some of these enzymes have "proof-reading" mechanisms. Here, an enzyme such asDNA polymerase catalyzes a reaction in a first step and then checks that the product is correct in a second step.[34] This two-step process results in average error rates of less than 1 error in 100 million reactions in high-fidelity mammalian polymerases.[1]: 5.3.1 Similar proofreading mechanisms are also found inRNA polymerase,[35]aminoacyl tRNA synthetases[36] andribosomes.[37]
Conversely, some enzymes displayenzyme promiscuity, having broad specificity and acting on a range of different physiologically relevant substrates. Many enzymes possess small side activities which arose fortuitously (i.e.neutrally), which may be the starting point for the evolutionary selection of a new function.[38][39]
Enzyme changes shape by induced fit upon substrate binding to form enzyme-substrate complex.Hexokinase has a large induced fit motion that closes over the substratesadenosine triphosphate andxylose. Binding sites in blue, substrates in black andMg2+ cofactor in yellow. (PDB:2E2N,2E2Q)
"Lock and key" model
To explain the observed specificity of enzymes, in 1894Emil Fischer proposed that both the enzyme and the substrate possess specific complementary geometric shapes that fit exactly into one another.[40] This is often referred to as "the lock and key" model.[1]: 8.3.2 This early model explains enzyme specificity, but fails to explain the stabilization of the transition state that enzymes achieve.[41]
Induced fit model
In 1958,Daniel Koshland suggested a modification to the lock and key model: since enzymes are rather flexible structures, the active site is continuously reshaped by interactions with the substrate as the substrate interacts with the enzyme.[42] As a result, the substrate does not simply bind to a rigid active site; the amino acidside-chains that make up the active site are molded into the precise positions that enable the enzyme to perform its catalytic function. In some cases, such asglycosidases, the substratemolecule also changes shape slightly as it enters the active site.[43] The active site continues to change until the substrate is completely bound, at which point the final shape and charge distribution is determined.[44]Induced fit may enhance the fidelity of molecular recognition in the presence of competition and noise via theconformational proofreading mechanism.[45]
Creating an environment with a charge distribution complementary to that of the transition state to lower its energy[47]
By providing an alternative reaction pathway:
Temporarily reacting with the substrate, forming a covalent intermediate to provide a lower energy transition state[48]
By destabilizing the substrate ground state:
Distorting bound substrate(s) into their transition state form to reduce the energy required to reach the transition state[49]
By orienting the substrates into a productive arrangement to reduce the reactionentropy change[50] (the contribution of this mechanism to catalysis is relatively small)[51]
Enzymes may use several of these mechanisms simultaneously. For example,proteases such astrypsin perform covalent catalysis using acatalytic triad, stabilize charge build-up on the transition states using anoxyanion hole, completehydrolysis using an oriented water substrate.[52]
Enzymes are not rigid, static structures; instead they have complex internal dynamic motions – that is, movements of parts of the enzyme's structure such as individual amino acid residues, groups of residues forming aprotein loop or unit ofsecondary structure, or even an entireprotein domain. These motions give rise to aconformational ensemble of slightly different structures that interconvert with one another atequilibrium. Different states within this ensemble may be associated with different aspects of an enzyme's function. For example, different conformations of the enzymedihydrofolate reductase are associated with the substrate binding, catalysis, cofactor release, and product release steps of the catalytic cycle,[53] consistent withcatalytic resonance theory. The transitions between the different conformations during the catalytic cycle involve internalviscoelatic motion that is facilitated by high-strain regions where amino acids are rearranged.[54]
Substrate presentation
Substrate presentation is a process where the enzyme is sequestered away from its substrate. Enzymes can be sequestered to the plasma membrane away from a substrate in the nucleus or cytosol.[55] Or within the membrane, an enzyme can be sequestered into lipid rafts away from its substrate in the disordered region. When the enzyme is released it mixes with its substrate. Alternatively, the enzyme can be sequestered near its substrate to activate the enzyme. For example, the enzyme can be soluble and upon activation bind to a lipid in the plasma membrane and then act upon molecules in the plasma membrane.[56]
Allosteric sites are pockets on the enzyme, distinct from the active site, that bind to molecules in the cellular environment. These molecules then cause a change in the conformation or dynamics of the enzyme that is transduced to the active site and thus affects the reaction rate of the enzyme.[57] In this way, allosteric interactions can either inhibit or activate enzymes. Allosteric interactions with metabolites upstream or downstream in an enzyme's metabolic pathway causefeedback regulation, altering the activity of the enzyme according to theflux through the rest of the pathway.[58]
Some enzymes do not need additional components to show full activity. Others require non-protein molecules called cofactors to be bound for activity.[59] Cofactors can be eitherinorganic (e.g., metalions andiron–sulfur clusters) ororganic compounds (e.g.,flavin andheme). These cofactors serve many purposes; for instance, metal ions can help in stabilizing nucleophilic species within the active site.[60] Organic cofactors can be eithercoenzymes, which are released from the enzyme's active site during the reaction, orprosthetic groups, which are tightly bound to an enzyme. Organic prosthetic groups can be covalently bound (e.g.,biotin in enzymes such aspyruvate carboxylase).[61]
An example of an enzyme that contains a cofactor iscarbonic anhydrase, which uses a zinc cofactor bound as part of its active site.[62] These tightly bound ions or molecules are usually found in the active site and are involved in catalysis.[1]: 8.1.1 For example, flavin and heme cofactors are often involved inredox reactions.[1]: 17
Enzymes that require a cofactor but do not have one bound are calledapoenzymes orapoproteins. An enzyme together with the cofactor(s) required for activity is called aholoenzyme (or haloenzyme). The termholoenzyme can also be applied to enzymes that contain multiple protein subunits, such as theDNA polymerases; here the holoenzyme is the complete complex containing all the subunits needed for activity.[1]: 8.1.1
Coenzymes
Coenzymes are small organic molecules that can be loosely or tightly bound to an enzyme. Coenzymes transport chemical groups from one enzyme to another.[63] Examples includeNADH,NADPH andadenosine triphosphate (ATP). Some coenzymes, such asflavin mononucleotide (FMN),flavin adenine dinucleotide (FAD),thiamine pyrophosphate (TPP), andtetrahydrofolate (THF), are derived fromvitamins. These coenzymes cannot be synthesized by the bodyde novo and closely related compounds (vitamins) must be acquired from the diet. The chemical groups carried include:
Since coenzymes are chemically changed as a consequence of enzyme action, it is useful to consider coenzymes to be a special class of substrates, or second substrates, which are common to many different enzymes. For example, about 1000 enzymes are known to use the coenzyme NADH.[64]
Coenzymes are usually continuously regenerated and their concentrations maintained at a steady level inside the cell. For example, NADPH is regenerated through thepentose phosphate pathway andS-adenosylmethionine bymethionine adenosyltransferase. This continuous regeneration means that small amounts of coenzymes can be used very intensively. For example, the human body turns over its own weight in ATP each day.[65]
Thermodynamics
The energies of the stages of achemical reaction. Uncatalysed (dashed line), substrates need a lot ofactivation energy to reach atransition state, which then decays into lower-energy products. When enzyme catalysed (solid line), the enzyme binds the substrates (ES), then stabilizes the transition state (ES‡) to reduce the activation energy required to produce products (EP) which are finally released.
As with all catalysts, enzymes do not alter the position of the chemical equilibrium of the reaction. In the presence of an enzyme, the reaction runs in the same direction as it would without the enzyme, just more quickly.[1]: 8.2.3 For example,carbonic anhydrase catalyzes its reaction in either direction depending on the concentration of its reactants:[66]
The rate of a reaction is dependent on theactivation energy needed to form thetransition state which then decays into products. Enzymes increase reaction rates by lowering the energy of the transition state. First, binding forms a low energy enzyme-substrate complex (ES). Second, the enzyme stabilises the transition state such that it requires less energy to achieve compared to the uncatalyzed reaction (ES‡). Finally the enzyme-product complex (EP) dissociates to release the products.[1]: 8.3
Enzymes can couple two or more reactions, so that a thermodynamically favorable reaction can be used to "drive" a thermodynamically unfavourable one so that the combined energy of the products is lower than the substrates. For example, the hydrolysis ofATP is often used to drive other chemical reactions.[67]
Enzyme kinetics is the investigation of how enzymes bind substrates and turn them into products.[68] The rate data used in kinetic analyses are commonly obtained fromenzyme assays. In 1913Leonor Michaelis andMaud Leonora Menten proposed a quantitative theory of enzyme kinetics, which is referred to asMichaelis–Menten kinetics.[69] The major contribution of Michaelis and Menten was to think of enzyme reactions in two stages. In the first, the substrate binds reversibly to the enzyme, forming the enzyme-substrate complex. This is sometimes called the Michaelis–Menten complex in their honor. The enzyme then catalyzes the chemical step in the reaction and releases the product. This work was further developed byG. E. Briggs andJ. B. S. Haldane, who derived kinetic equations that are still widely used today.[70]
Enzyme rates depend onsolution conditions and substrateconcentration. To find the maximum speed of an enzymatic reaction, the substrate concentration is increased until a constant rate of product formation is seen. This is shown in the saturation curve on the right. Saturation happens because, as substrate concentration increases, more and more of the free enzyme is converted into the substrate-bound ES complex. At the maximum reaction rate (Vmax) of the enzyme, all the enzyme active sites are bound to substrate, and the amount of ES complex is the same as the total amount of enzyme.[1]: 8.4
Vmax is only one of several important kinetic parameters. The amount of substrate needed to achieve a given rate of reaction is also important. This is given by theMichaelis–Menten constant (Km), which is the substrate concentration required for an enzyme to reach one-half its maximum reaction rate; generally, each enzyme has a characteristicKM for a given substrate. Another useful constant iskcat, also called theturnover number, which is the number of substrate molecules handled by one active site per second.[1]: 8.4
The efficiency of an enzyme can be expressed in terms ofkcat/Km. This is also called the specificity constant and incorporates therate constants for all steps in the reaction up to and including the first irreversible step. Because the specificity constant reflects both affinity and catalytic ability, it is useful for comparing different enzymes against each other, or the same enzyme with different substrates. The theoretical maximum for the specificity constant is called the diffusion limit and is about 108 to 109 (M−1 s−1). At this point every collision of the enzyme with its substrate will result in catalysis, and the rate of product formation is not limited by the reaction rate but by the diffusion rate. Enzymes with this property are calledcatalytically perfect orkinetically perfect. Example of such enzymes aretriose-phosphate isomerase,carbonic anhydrase,acetylcholinesterase,catalase,fumarase,β-lactamase, andsuperoxide dismutase.[1]: 8.4.2 The turnover of such enzymes can reach several million reactions per second.[1]: 9.2 But most enzymes are far from perfect: the average values of and are about and, respectively.[71]
Michaelis–Menten kinetics relies on thelaw of mass action, which is derived from the assumptions of freediffusion and thermodynamically driven random collision. Many biochemical or cellular processes deviate significantly from these conditions, because ofmacromolecular crowding and constrained molecular movement.[72] More recent, complex extensions of the model attempt to correct for these effects.[73]
Inhibition
The coenzymefolic acid (left) and the anti-cancer drugmethotrexate (right) are very similar in structure (differences show in green). As a result, methotrexate is a competitive inhibitor of many enzymes that use folates.
Enzyme reaction rates can be decreased by various types of enzyme inhibitors.[74]: 73–74
Types of inhibition
Competitive
Acompetitive inhibitor and substrate cannot bind to the enzyme at the same time.[75] Often competitive inhibitors strongly resemble the real substrate of the enzyme. For example, the drugmethotrexate is a competitive inhibitor of the enzymedihydrofolate reductase, which catalyzes the reduction ofdihydrofolate to tetrahydrofolate.[76] The similarity between the structures of dihydrofolate and this drug are shown in the accompanying figure. This type of inhibition can be overcome with high substrate concentration. In some cases, the inhibitor can bind to a site other than the binding-site of the usual substrate and exert anallosteric effect to change the shape of the usual binding-site.[77]
Non-competitive
Anon-competitive inhibitor binds to a site other than where the substrate binds. The substrate still binds with its usual affinity and hence Km remains the same. However the inhibitor reduces the catalytic efficiency of the enzyme so that Vmax is reduced. In contrast to competitive inhibition, non-competitive inhibition cannot be overcome with high substrate concentration.[74]: 76–78
Uncompetitive
Anuncompetitive inhibitor cannot bind to the free enzyme, only to the enzyme-substrate complex; hence, these types of inhibitors are most effective at high substrate concentration. In the presence of the inhibitor, the enzyme-substrate complex is inactive.[74]: 78 This type of inhibition is rare.[78]
Mixed
Amixed inhibitor binds to an allosteric site and the binding of the substrate and the inhibitor affect each other. The enzyme's function is reduced but not eliminated when bound to the inhibitor. This type of inhibitor does not follow the Michaelis–Menten equation.[74]: 76–78
In many organisms, inhibitors may act as part of afeedback mechanism. If an enzyme produces too much of one substance in the organism, that substance may act as an inhibitor for the enzyme at the beginning of the pathway that produces it, causing production of the substance to slow down or stop when there is sufficient amount. This is a form ofnegative feedback. Major metabolic pathways such as thecitric acid cycle make use of this mechanism.[1]: 17.2.2
Since inhibitors modulate the function of enzymes they are often used as drugs. Many such drugs are reversible competitive inhibitors that resemble the enzyme's native substrate, similar tomethotrexate above; other well-known examples includestatins used to treat highcholesterol,[82] andprotease inhibitors used to treatretroviral infections such asHIV.[83] A common example of an irreversible inhibitor that is used as a drug isaspirin, which inhibits theCOX-1 andCOX-2 enzymes that produce theinflammation messengerprostaglandin.[81] Other enzyme inhibitors are poisons. For example, the poisoncyanide is an irreversible enzyme inhibitor that combines with the copper and iron in the active site of the enzymecytochrome c oxidase and blockscellular respiration.[84]
Factors affecting enzyme activity
As enzymes are made up of proteins, their actions are sensitive to change in many physio chemical factors such as pH, temperature, substrate concentration, etc.
The following table shows pH optima for various enzymes.[85]
An important function of enzymes is in thedigestive systems of animals. Enzymes such asamylases andproteases break down large molecules (starch orproteins, respectively) into smaller ones, so they can be absorbed by the intestines. Starch molecules, for example, are too large to be absorbed from the intestine, but enzymes hydrolyze the starch chains into smaller molecules such asmaltose and eventuallyglucose, which can then be absorbed. Different enzymes digest different food substances. Inruminants, which haveherbivorous diets, microorganisms in the gut produce another enzyme,cellulase, to break down the cellulose cell walls of plant fiber.[90]
Metabolism
Themetabolic pathway ofglycolysis releases energy by convertingglucose topyruvate via a series of intermediate metabolites. Each chemical modification (red box) is performed by a different enzyme.
Several enzymes can work together in a specific order, creatingmetabolic pathways.[1]: 30.1 In a metabolic pathway, one enzyme takes the product of another enzyme as a substrate. After the catalytic reaction, the product is then passed on to another enzyme. Sometimes more than one enzyme can catalyze the same reaction in parallel; this can allow more complex regulation: with, for example, a low constant activity provided by one enzyme but an inducible high activity from a second enzyme.[91]
Enzymes determine what steps occur in these pathways. Without enzymes, metabolism would neither progress through the same steps and could not be regulated to serve the needs of the cell. Most central metabolic pathways are regulated at a few key steps, typically through enzymes whose activity involves the hydrolysis of ATP. Because this reaction releases so much energy, other reactions that arethermodynamically unfavorable can be coupled to ATP hydrolysis, driving the overall series of linked metabolic reactions.[1]: 30.1
Control of activity
There are five main ways that enzyme activity is controlled in the cell.[1]: 30.1.1
Regulation
Enzymes can be eitheractivated orinhibited by other molecules. For example, the end product(s) of a metabolic pathway are often inhibitors for one of the first enzymes of the pathway (usually the first irreversible step, called committed step), thus regulating the amount of end product made by the pathways. Such a regulatory mechanism is called anegative feedback mechanism, because the amount of the end product produced is regulated by its own concentration.[92]: 141–48 Negative feedback mechanism can effectively adjust the rate of synthesis of intermediate metabolites according to the demands of the cells. This helps with effective allocations of materials and energy economy, and it prevents the excess manufacture of end products. Like otherhomeostatic devices, the control of enzymatic action helps to maintain a stable internal environment in living organisms.[92]: 141
Post-translational modification
Examples ofpost-translational modification includephosphorylation,myristoylation andglycosylation.[92]: 149–69 For example, in the response toinsulin, thephosphorylation of multiple enzymes, includingglycogen synthase, helps control the synthesis or degradation ofglycogen and allows the cell to respond to changes inblood sugar.[93] Another example of post-translational modification is the cleavage of the polypeptide chain.Chymotrypsin, a digestive protease, is produced in inactive form aschymotrypsinogen in thepancreas and transported in this form to thestomach where it is activated. This stops the enzyme from digesting the pancreas or other tissues before it enters the gut. This type of inactive precursor to an enzyme is known as azymogen[92]: 149–53 or proenzyme.
Enzymes can be compartmentalized, with different metabolic pathways occurring in differentcellular compartments. For example,fatty acids are synthesized by one set of enzymes in thecytosol,endoplasmic reticulum andGolgi and used by a different set of enzymes as a source of energy in themitochondrion, throughβ-oxidation.[96] In addition,trafficking of the enzyme to different compartments may change the degree ofprotonation (e.g., the neutralcytoplasm and the acidiclysosome) or oxidative state (e.g., oxidizingperiplasm or reducingcytoplasm) which in turn affects enzyme activity.[97] In contrast to partitioning into membrane bound organelles, enzyme subcellular localisation may also be altered through polymerisation of enzymes into macromolecular cytoplasmic filaments.[98][99]
Organ specialization
Inmulticellulareukaryotes, cells in differentorgans andtissues have different patterns ofgene expression and therefore have different sets of enzymes (known asisozymes) available for metabolic reactions. This provides a mechanism for regulating the overall metabolism of the organism. For example,hexokinase, the first enzyme in theglycolysis pathway, has a specialized form calledglucokinase expressed in the liver andpancreas that has a loweraffinity for glucose yet is more sensitive to glucose concentration.[100] This enzyme is involved in sensingblood sugar and regulating insulin production.[101]
Involvement in disease
Inphenylalanine hydroxylase over 300 different mutations throughout the structure causephenylketonuria.Phenylalanine substrate andtetrahydrobiopterin coenzyme in black, andFe2+ cofactor in yellow. (PDB:1KW0)Hereditary defects in enzymes are generally inherited in anautosomal fashion because there are more non-X chromosomes than X-chromosomes, and arecessive fashion because the enzymes from the unaffected genes are generally sufficient to prevent symptoms in carriers.
Since the tight control of enzyme activity is essential forhomeostasis, any malfunction (mutation, overproduction, underproduction or deletion) of a single critical enzyme can lead to a genetic disease. The malfunction of just one type of enzyme out of the thousands of types present in the human body can be fatal. An example of a fatal genetic disease due to enzyme insufficiency isTay–Sachs disease, in which patients lack the enzymehexosaminidase.[102][103]
One example of enzyme deficiency is the most common type ofphenylketonuria. Many different single amino acid mutations in the enzymephenylalanine hydroxylase, which catalyzes the first step in the degradation ofphenylalanine, result in build-up of phenylalanine and related products. Some mutations are in the active site, directly disrupting binding and catalysis, but many are far from the active site and reduce activity by destabilising the protein structure, or affecting correct oligomerisation.[104][105] This can lead tointellectual disability if the disease is untreated.[106] Another example ispseudocholinesterase deficiency, in which the body's ability to break down choline ester drugs is impaired.[107]Oral administration of enzymes can be used to treat some functional enzyme deficiencies, such aspancreatic insufficiency[108] andlactose intolerance.[109]
Similar to any other protein, enzymes change over time throughmutations and sequence divergence. Given their central role inmetabolism, enzyme evolution plays a critical role inadaptation. A key question is therefore whether and how enzymes can change their enzymatic activities alongside. It is generally accepted that many new enzyme activities have evolved throughgene duplication and mutation of the duplicate copies although evolution can also happen without duplication. One example of an enzyme that has changed its activity is the ancestor ofmethionyl aminopeptidase (MAP) and creatine amidinohydrolase (creatinase) which are clearly homologous but catalyze very different reactions (MAP removes the amino-terminalmethionine in new proteins while creatinase hydrolysescreatine tosarcosine andurea). In addition, MAP is metal-ion dependent while creatinase is not, hence this property was also lost over time.[112] Small changes of enzymatic activity are extremely common among enzymes. In particular, substrate binding specificity (see above) can easily and quickly change with single amino acid changes in their substrate binding pockets. This is frequently seen in the main enzyme classes such askinases.[113]
Artificial (in vitro) evolution is now commonly used to modify enzyme activity or specificity for industrial applications (see below).
Enzymes are used in thechemical industry and other industrial applications when extremely specific catalysts are required. Enzymes in general are limited in the number of reactions they have evolved to catalyze and also by their lack of stability inorganic solvents and at high temperatures. As a consequence,protein engineering is an active area of research and involves attempts to create new enzymes with novel properties, either through rational design orin vitro evolution.[114][115] These efforts have begun to be successful, and a few enzymes have now been designed "from scratch" to catalyze reactions that do not occur in nature.[116]
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Further reading
General
Berg JM, Tymoczko JL, Stryer L (2002).Biochemistry (5th ed.). New York, NY: W. H. Freeman.ISBN0-7167-3051-0., A biochemistry textbook available free online through NCBI Bookshelf.
