Enzyme

From New World Encyclopedia
Ribbon diagram of the enzyme TIM. TIM is catalytically perfect, meaning its conversion rate is limited, or nearly limited to its substrate diffusion rate.

An enzyme is a protein that catalyzes, or speeds up, a chemical reaction. The word comes from the Greek ένζυμο, énsymo, which comes from én ("at" or "in") and simo ("leaven" or "yeast"). Certain RNAs also have catalytic activity, but to differentiate them from protein enzymes, they are referred to as RNA enzymes or ribozymes.

Enzymes are essential to sustain life because most chemical reactions in biological cells would occur too slowly, or would lead to different products without enzymes. A malfunction (mutation, overproduction, underproduction or deletion) of a single critical enzyme can lead to a severe disease. For example, the most common type of phenylketonuria is caused by a single amino acid mutation in the enzyme phenylalanine hydroxylase, which catalyzes the first step in the degradation of phenylalanine. The resulting build-up of phenylalanine and related products can lead to mental retardation if the disease is untreated.

Like all catalysts, enzymes work by providing an alternate pathway of lower activation energy of a reaction, thus allowing the reaction to proceed much faster. Enzymes may speed up reactions by a factor of many millions. An enzyme, like any catalyst, remains unaltered by the completed reaction and can therefore continue to function. Because enzymes do not affect the relative energy between the products and reagents, they do not affect equilibrium of a reaction. However, the advantage of enzymes compared to most other catalysts is their sterio-, regio- and chemoselectivity and specificity.

Enzyme activity can be affected by other molecules. Inhibitors are naturally occurring or synthetic molecules that decrease or abolish enzyme activity; activators are molecules that increase activity. Some irreversible inhibitors bind enzymes very tightly, effectively inactivating them. Many drugs and poisons act by inhibiting enzymes. Aspirin inhibits the COX-1 and COX-2 enzymes that produce the inflammation messenger prostaglandin, thus suppressing pain and inflammation. The poison cyanide inhibits cytochrome c oxidase, which effectively blocks cellular respiration.

While all enzymes have a biological role, some enzymes are used commercially for other purposes. Many household cleaners use enzymes to speed up chemical reactions ( e.g., breaking down protein or starch stains in clothes).

More than 5,000 enzymes are known. Typically the suffix -ase is added to the name of the substrate (e.g., lactase is the enzyme that catalyzes the cleavage of lactose) or the type of reaction (e.g., DNA polymerase catalyzes the formation of DNA polymers). However, this is not always the case, especially when enzymes modify multiple substrates. For this reason Enzyme Commission or EC numbers are used to classify enzymes based on the reactions they catalyze. Even this is not a perfect solution, as enzymes from different species or even very similar enzymes in the same species may have identical EC numbers.

Etymology and history

Eduard Buchner

The word enzyme comes from Greek: "in leaven". As early as the late 1700s and early 1800s, the digestion of meat by stomach secretions and the conversion of starch to sugars by plant extracts and saliva were observed.

Studying the fermentation of sugar to alcohol by yeast, Louis Pasteur came to the conclusion that this fermentation was catalyzed by "ferments" in the yeast, which were thought to function only in the presence of living organisms.

In 1897, Hans and Eduard Buchner inadvertently used yeast extracts to ferment sugar, despite the absence of living yeast cells. They were interested in making extracts of yeast cells for medical purposes, and, as one possible way of preserving them, they added large amounts of sucrose to the extract. To their surprise, they found that the sugar was fermented, even though there were no living yeast cells in the mixture. The term "enzyme" was used to describe the substance(s) in yeast extract that brought about the fermentation of sucrose. It was not until 1926, however, that the first enzyme was obtained in pure form.

The structure of enzymes

Enzyme structure is important because it determines the enzyme's particular function in the body. Enzymes (and other proteins) are composed of amino acid chains; the linear sequence of amino acids determines the characteristic folding of the chains into a three-dimensional structure. An enzyme might contain only one polypeptide chain, typically linking one hundred or more amino acids, or it might consist of several polypeptide chains that act together as a unit.

File:Enzymeactivesite.png
The active site of an enzyme.

Most enzymes are larger than the substrates on which they act. Only a very small portion of the enzyme, approximately 10 amino acids, comes into direct contact with the substrate(s). This region, where the binding of the substrate(s) and the reaction occur, is known as the active site of the enzyme.

Specificity

Enzymes are usually specific as to the reactions they catalyze and the substrates that are involved in these reactions. An enzyme combines with its substrate(s) to form a short-lived enzyme-substrate complex. There are two models to explain how the binding of enzyme and substrate occurs: "lock and key" and induced fit.

"Lock and key" model

The induced fit model of enzyme action.

To account for the specificity of enzymes, Emil Fischer proposed that the enzyme had a particular shape into which the substrate(s) fit exactly. This model of exact fit, introduced in the 1890s, is often referred to as the "lock and key" model.

Induced fit model

In 1958, Daniel Koshland suggested a modification to the "lock and key" model. Enzymes are rather flexible structures. The active site of an enzyme can be modified as the substrate interacts with the enzyme, creating an "induced fit" between enzyme and substrate. The amino acids side chains that make up the active site are molded into a precise shape, which enables the enzyme to perform its catalytic function. In some cases, the substrate molecule changes shape slightly as it enters the active site.

Enzyme cofactors

Some enzymes do not need any additional components to exhibit full activity. However, others require non-protein molecules to be bound for activity. Cofactors can be either inorganic (e.g., metal ions and Iron-sulfur clusters) or organic compounds, which are also known as coenzymes.

Enzymes that require a cofactor, but do not have one bound are called apoenzymes. An apoenzyme together with its cofactor(s) constitutes a holoenzyme (i.e, the active form). Most cofactors are not covalently bound to an enzyme, but are closely associated. However, some cofactors known as prosthetic groups are covalently bound (e.g., thiamine pyrophosphate in certain enzymes).

Most cofactors are either regenerated or chemically unchanged at the end of the reactions. Many cofactors are vitamin-derivatives and serve as carriers to transfer electrons, atoms, or functional groups from an enzyme to a substrate. Common examples are NAD and NADP, which are involved in electron transfer and coenzyme A, which is involved in the transfer of acetyl groups.

How enzymes catalyze reactions

Diagram of a catalytic reaction, showing the energy niveau at each stage of the reaction. The substrates usually need a large amount of energy to reach the transition state, which then reacts to form the end product. The enzyme stabilizes the transition state, reducing the energy of the transition state and thus the energy required to get over this barrier.

As with all catalysts, all reactions catalyzed by enzymes must be "spontaneous" (containing a net negative Gibbs free energy). With the enzyme, they run in the same direction as they would without the enzyme, just more quickly. However, the uncatalyzed, "spontaneous" reaction might lead to different products than the catalyzed reaction. Furthermore, enzymes can couple two or more reactions, so that a thermodynamically favorable reaction can be used to "drive" a thermodynamically unfavorable one. For example, the cleavage of the high-energy compound ATP is often used to drive other, energetically unfavorable chemical reactions.

Enzymes catalyze the forward and backward reactions equally. They do not alter the equilibrium itself, but only the speed at which it is reached. Carbonic anhydrase catalyzes its reaction in either direction depending on the conditions.

(in tissues - high CO2 concentration)
(in lungs - low CO2 concentration)

Kinetics

In 1913, Leonor Michaelis and Maud Menten proposed a quantitative theory of enzyme kinetics, which is referred to as Michaelis-Menten kinetics. Their work was further developed by G. E. Briggs and J. B. S. Haldane, who derived numerous kinetic equations that are still widely used today.

Enzymes can perform up to several million catalytic reactions per second; to determine the maximum speed of an enzymatic reaction, the substrate concentration is increased until a constant rate of product formation is achieved. This is the maximum velocity (Vmax) of the enzyme. In this state, all enzyme active sites are saturated with substrate. However, Vmax is only one kinetic parameter that biochemists are interested in. The amount of substrate needed to achieve a given rate of reaction is also of interest. This can be expressed by the Michaelis-Menten constant (Km), which is the substrate concentration required for an enzyme to reach one half its maximum velocity. Each enzyme has a characteristic Km for a given substrate.

The efficiency of an enzyme can be expressed in terms of kcat/Km. The quantity kcat, also called the turnover number, incorporates the rate constants for all steps in the reaction, and is the quotient of Vmax and the total enzyme concentration. kcat/Km is a useful quantity for comparing different enzymes against each other, or the same enzyme with different substrates, because it takes both affinity and catalytic ability into consideration. The theoretical maximum for kcat/Km, called the diffusion limit, 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 that reach this kcat/Km value are called catalytically perfect or kinetically perfect. Example of such enzymes are triose-phosphate isomerase, carbonic anhydrase, acetylcholinesterase, catalase, fumarase, ß-lactamase, and superoxide dismutase.

Some enzymes operate with kinetics which are faster than diffusion rates, which would seem to be impossible. Several mechanisms have been invoked to explain this phenomenon. Some proteins are believed to accelerate catalysis by drawing their substrate in and preorienting them by using dipolar electric fields. Some invoke a quantum-mechanical tunneling explanation whereby a proton or an electron can tunnel through activation barriers, although for protons tunneling remains somewhat controversial. [1][2]

Regulation of enzyme activity

A competitive inhibitor binds reversibly to the active site of the enzyme, preventing the binding of substrate.
Diagram showing the mechanism of non-competitive inhibition.

Compounds called inhibitors can decrease enzyme reaction rates through competitive or non-competitive inhibition.

Competitive inhibition

In competitive inhibition, the inhibitor binds to the substrate binding site as shown (right part b), thus preventing substrate binding.


Non-competitive inhibition

Non-competitive inhibitors never bind to the active site, but to other parts of the enzyme that can be far away from the substrate binding site, consequently, there is no competition between the substrate and inhibitor for the enzyme. The extent of inhibition depends entirely on the inhibitor concentration and will not be affected by the substrate concentration. For example, cyanide combines with the copper prosthetic groups of the enzyme cytochrome c oxidase, thus inhibiting cellular respiration. This type of inhibition is typically irreversible, meaning that the enzyme will no longer function.

By changing the conformation (the three-dimensional structure) of the enzyme, the inhibitors either disable the ability of the enzyme to bind or turn over its substrate. The enzyme-inhibitor (EI) and enzyme-inhibitor-substrate (EIS) complex have no catalytic activity.

Metabolic pathways and allosteric enzymes

Several enzymes can work together in a specific order, creating metabolic pathways. 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. The end product(s) of such a 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 a negative feedback mechanism, because the amount of the end product produced is regulated by its own concentration. 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 other homeostatic devices, the control of enzymatic action helps to maintain a stable internal environment in living organisms.

Enzyme-naming conventions

By common convention, an enzyme's name consists of a description of what it does, with the word ending in -ase. Examples are alcohol dehydrogenase and DNA polymerase. Kinases are enzymes that transfer phosphate groups. This results in different enzymes with the same function having the same basic name; they are therefore distinguished by other characteristics, such as their optimal pH (alkaline phosphatase) or their location (membrane ATPase). Furthermore, the reversibility of chemical reactions means that the normal physiological direction of an enzyme's function may not be that observed under laboratory conditions. This can result in the same enzyme being identified with two different names: one stemming from the formal laboratory identification as described above, the other representing its behavior in the cell. For instance the enzyme formally known as xylitol:NAD+ 2-oxidoreductase (D-xylulose-forming) is more commonly referred to in the cellular physiological sense as D-xylulose reductase, reflecting the fact that the function of the enzyme in the cell is actually the reverse of what is often seen under in vitro conditions.

The International Union of Biochemistry and Molecular Biology has developed a nomenclature for enzymes, the EC numbers; each enzyme is described by a sequence of four numbers, preceded by "EC". The first number broadly classifies the enzyme based on its mechanism:

The toplevel classification is

  • EC 1 Oxidoreductases: catalyze oxidation/reduction reactions
  • EC 2 Transferases: transfer a functional group (e.g. a methyl or phosphate group)
  • EC 3 Hydrolases: catalyze the hydrolysis of various bonds
  • EC 4 Lyases: cleave various bonds by means other than hydrolysis and oxidation
  • EC 5 Isomerases: catalyze isomerization changes within a single molecule
  • EC 6 Ligases: join two molecules with covalent bonds

The complete nomenclature can be browsed at http://www.chem.qmul.ac.uk/iubmb/enzyme/

Industrial Applications

Application

Enzymes used

Uses

Notes and examples

Biological detergent Primarily proteases, produced in an extracellular form from bacteria Used for presoak conditions and direct liquid applications helping with removal of protein stains from clothes.
Amylase enzymes Detergents for machine dish washing to remove resistant starch residues.
Baking industry Fungal alpha-amylase enzymes: normally inactivates about 50 degrees Celsius, destroyed during baking process Catalyze breakdown of starch in the flour to sugar. Yeast action on sugar produces carbon dioxide. Used in production of white bread, buns, and rolls
File:Amylose.gif
alpha-amylase catalyzes the release of sugar monomers from starch
Protease enzymes Biscuit manufacturers use them to lower the protein level of flour.
Baby foods Trypsin To predigest baby foods
Brewing industry Enzymes from barley are released during the mashing stage of beer production. They degrade starch and proteins to produce simple sugar, amino acids and peptides that are used by yeast to enhance fermentation.
Germinating barley used for malt.
Industrially produced barley enzymes. Widely used in the brewing process to substitute for the natural enzymes found in barley.
Amylase, glucanases, proteases Split polysaccharides and proteins in the malt
Betaglucosidase Improve the filtration characteristics.
Amyloglucosidase Low-calorie beer
Proteases Remove cloudiness during storage of beers.
Fruit juices Cellulases, pectinases Clarify fruit juices
Dairy industry Rennin, derived from the stomachs of young ruminant animals (calves, lambs) Manufacture of cheese, used to split protein Note: As animals age rennin production decreases and is replaced by another protease, pepsin, which is not suitable for cheese production. In recent years the increase in cheese consumption, as well as increased beef production, has resulted in a shortage of rennin and escalating prices.
Microbially produced enzyme Now finding increasing use in the dairy industry
File:Roquefort cheese.jpg
Roquefort cheese
Lipases Is implemented during the production of Roquefort cheese to enhance the ripening of the blue-mould cheese.
Lactases Break down lactose to glucose and galactose
Starch industry Amylases, amyloglucosideases and glucoamylases Converts starch into glucose and various syrups
Glucose isomerase Converts glucose into fructose (high fructose syrups derived from starchy materials have enhanced sweetening properties and lower calorific values)
Rubber industry Catalase To generate oxygen from peroxide to convert latex to foam rubber
Paper industry Amylases Degrade starch to lower viscosity product needed for sizing and coating paper
Photographic industry Protease (ficin) Dissolve gelatin off the scrap film allowing recovery of silver present


References
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  • Koshland D. The Enzymes, v. I, ch. 7, Acad. Press, New York, 1959
  • Perutz M. Proc. Roy. Soc., B (1967) 167, 448,
  • Cha, Y., Murray, C. J. & Klinman, J. P. Science (1989) 243, 1325-1330 .
  • Leonor Michaelis and Maud Menten, Die Kinetik der Invertinwirkung, Biochem. Z. (1913) 49, 333-369.
  • G. E. Briggs and J. B. S. Haldane, A note on the kinetics of enzyme action, Biochem. J., (1925) 19, 339-339.
  • M.V. Volkenshtein, R.R. Dogonadze, A.K. Madumarov, Z.D. Urushadze, Yu.I. Kharkats. Theory of Enzyme Catalysis.- Molekuliarnaya Biologia, (1972), 431-439 (In Russian, English summary)

External links

  • ExPASy enzyme database, links to Swiss-Prot sequence data, entries in other databases and to related literature searches
  • PDBsum links to the known 3-D structure data of enzymes in the Protein Data Bank
  • MACiE, database of enzyme reaction mechanisms
  • BRENDA, comprehensive compilation of information and literature references about all known enzymes; requires payment by commercial users
  • Weizmann Institute's Genecards Database, extensive database of protein properties and their associated genes.
  • Cytochrome P450 enzymes site lists over 4000 versions of enzymes from this cytochrome in plants and animals

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  1. Mireia Garcia-Viloca,1 Jiali Gao,1 Martin Karplus,2* Donald G. Truhlar Science 9 January 2004: Vol. 303. no. 5655, pp. 186 - 195
  2. Olsson MH, Siegbahn PE, Warshel A. J Am Chem Soc. 2004 Mar 10;126(9):2820-8.