Enzyme

Etymology and history

]] The word comes from Greek language|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 "Vitalism|ferments" in the yeast, which were thought to function only in the presence of living organisms. In 1897, Hans Buchner|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.

3D-Structure

In enzymes, as with other proteins, function is determined by structure. An enzyme can be:

A monomer|monomeric protein, ''i.e.'', containing only one polypeptide chain, typically one hundred or more amino acids; or

an oligomeric protein consisting of several polypeptide chains, different or identical, that act together as a unit. As with any protein, each monomer is actually produced as a long, linear chain of amino acids, which folds in a particular fashion to produce a three-dimensional product. Individual monomers may then combine via non-covalent interactions to form a multimeric protein. Most enzymes are far larger molecules than the substrates they act on and that only a very small portion of the enzyme, around 10 amino acids, come into direct contact with the substrate(s). This region, where binding of the substrate(s) and then the reaction occurs, is known as the active site of the enzyme. Some enzymes contain sites thst bind cofactors, which are needed for catalysis. Certain enzymes have binding sites for small molecules, which are often direct or #Metabolic pathways|indirect products or substrates of the reaction catalyzed. This binding can serve to increase or decrease the enzyme's activity (depending on the molecule and enzyme), providing a means for feedback regulation.

Specificity

Enzymes are usually specific as to the reactions they catalyze and the substrate (biochemistry)|substrates that are involved in these reactions. Shape, charge complementarity, and hydrophillic/hydrophobic character of enzyme and substrate are responsible for this specificity.

"Lock and key" hypothesis

Enzymes are very specific and it was suggested by Emil Fischer in 1890 that this was because the enzyme had a particular shape into which the substrate(s) fit exactly. This is often referred to as "the lock and key" hypothesis. An enzyme combines with its substrate(s) to form a short-lived enzyme-substrate complex.

Induced fit hypothesis

In 1958 Daniel Koshland suggested a modification to the "lock and key" hypothesis. Enzymes are rather flexible structures. The active site of an enzyme could be modified as the substrate interacts with the enzyme. The amino acids sidechains which 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. A suitable analogy would be that of a hand changing the shape of a glove as the glove is put on.

Modifications

Many enzymes contain not only a protein part but need additionally various modifications. These modifications are made ''posttranslational'', ''i.e.'', after the polypeptide chain was synthesized. Additional groups can be synthesized onto the polypeptide chain, ''e.g.'' phosphorylation or glycosylation of the enzyme. Another kind of posttranslational modification is the cleavage and splicing of the polypeptide chain. Chymotrypsin, a digestive protease, is produced in inactive form as chymotrypsinogen in the pancreas and transported in this form to the stomach where it is activated. This prevents the enzyme from harmful digestion of the pancreas or other tissue. This type of inactive precursor to an enzyme is known as a zymogen.

Enzyme cofactors

Some enzymes do not need any additional components to exhibit full activities. 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 molecules|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.,'' heme in hemoglobin. 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 Nicotinamide adenine dinucleotide|NAD and NADP, which are involved in electron transfer and Coenzyme A, which is involved in the transfer of acetyl groups.

Allosteric modulation

Allosteric enzymes have either effector binding sites, or multiple protein subunits that interact with each other and thus influence catalytic activity.

Kinetics

In 1913, Leonor Michaelis and Maud Menten proposed a quantitative theory of enzyme kinetics which is still widely used today (usually referred to as Michaelis-Menten kinetics). 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 (''V''_max) of the enzyme. In this state, all enzyme active sites are saturated with substrate. However, ''V''_max 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 (''K''_m), which is the substrate concentration required for an enzyme to reach one half its maximum velocity. Each enzyme has a characteristic ''K''_m for a given substrate. Since ''V''_max cannot be measured directly, both ''K''_m and ''V''_max are usually determined by extrapolating from a limited data set, using what is known as a double reciprocal, or Lineweaver-Burk plot. The efficiency of an enzyme can be expressed in terms of ''k''_cat/''K''_m. The quantity ''k''_cat, also called the turnover number, incorporates the rate constants for all steps in the reaction, and is the quotient of ''V''_max and the total enzyme concentration. ''k''_cat/''K''_m 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 ''k''_cat/''K''_m, called 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 ''k''_cat/''K''_m value are called ''catalytically perfect'' or ''kinetically perfect''. Example of such enzymes are triosephosphateisomerase|triose-phosphate isomerase, carbonic anhydrase, acetylcholinesterase, catalase, fumarase, ß-lactamase, and superoxide dismutase. The Quantum mechanics|quantum-mechanical (physical) model of enzyme catalysis explains how certain enzymes worked faster than previously thought possible. This is achived by a process known as Quantum tunneling|tunneling. While proposed in the early 1970s, it was not until 1989 that evidence of tunneling was found.

Thermodynamics

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 Adenosine triphosphate|ATP is often used to drive other, energetically unfavorable chemical reactions. Many reactions catalyzed by an enzyme are reversible. : $\backslash mathrm$ Enzymes catalyze the forward and backward reactions equally. They do not alter the equilibrium itself, but only the speed at which it is reached, for example, carbonic anhydrase which catalyzes a reaction in either direction depending on the conditions at the time. : $\backslash mathrm$ (in Biological tissue|tissues - high CO₂ concentration) : $\backslash mathrm$ (in lungs - low CO₂ concentration)

Inhibition

Enzymes reaction rates can be changed by competitive inhibition, non-competitive inhibition, uncompetitive inhibition and mixed inhibition.

Competitive inhibition

The inhibitor may bind to the substrate binding site as shown in the figure above, thus preventing substrate binding. An example for competitive inhibition is the enzyme succinate dehydrogenase by malonate. Succinate dehydrogenase catalyses the oxidation of succinate to fumarate. (bottom).]]

Uncompetitive inhibition

Uncompetitive inhibition occurs when the inhibitor binds only to the enzyme-substrate complex, not to the free enzyme, the enzyme-inhibitor-substrate (EIS) complex is catalytically inactive. This mode of inhibition is rare.

Non-competitive inhibition

Non-competitive inhibitors never bind to the active center, 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. However, these inhibitors bind only loosely with the enzyme and can be removed to resume the enzymatic activities. For example, cyanide combines with the copper prosthetic groups of the enzyme cytochrome c oxidase, thus inhibiting cellular respiration. By changing the Chemical conformation|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 EI and EIS-complex have no catalytic activity.

Partially competitive inhibition

The mechanism of partially competitive is similar to that of non-competitive inhibition, except that the EIS-complex has catalytic activity, which may be lower or even higher (partially competitive activation) than that of the ES-complex.

Irreversible inhibitors

Some inhibitors bind irreversibly with the enzyme molecules, inhibiting the catalytic activities permanently. The enzymatic reactions will stop sooner or later and are not affected by an increase in substrate concentration. These are irreversible inhibitors. Examples are heavy metal ions including silver (element)|silver, mercury (element)|mercury and lead (element)|lead ions. Another example of irreversible inhibition is provided by the nerve gas diisopropylfluorophosphate (DFP) designed for use in warfare. It combines with the amino acid serine (contains the —OH group) at the active site of the enzyme acetylcholinesterase. The enzyme deactivates the neurotransmitter acetylcholine. Neurotransmitters are needed to continue the passage of nerve impulses from one neuron to another across the synapse. Once the impulse has been transmitted, acetylcholinesterase functions to deactivate the acetycholine almost immediately by breaking it down. If the enzyme is inhibited, acetylcholine accumulates and nerve impulses cannot be stopped, causing prolonged muscle contraction. Paralysis occurs and death may result since the diaphragm|respiratory muscles are affected. Some insecticides currently in use, including those known as organophosphates (e.g. parathion), have a similar effect on insects, and can also cause harm to nervous system|nervous and muscular system of humans who are overexposed to them.

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|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 homeostasis|homeostatic devices, the control of enzymatic action helps to maintain a stable internal environment in living organisms. Enzymes that are regulated by end-production inhibition are usually allosteric enzymes. An allosteric enzyme molecule has an active site and also an allosteric site. The allosteric site can bind with allosteric effectors that affect the activity of the enzyme molecule. Allosteric effectors include allosteric activators and allosteric inhibitors. The binding with an allosteric activator activates an enzyme molecule because the active site is in the right conformation to bind with substrate molecules. The binding with an allosteric inhibitor inactivates the enzyme molecule because the conformation of the active site is altered. The activation and inhibition of an allosteric enzyme are reversible. , the enzyme of the first reaction in the pathway, is an allosteric enzyme, and CTP, the end product, is an allosteric inhibitor of ATCase.]]

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 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 http://www.iubmb.unibe.ch/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

See also

List of enzymes

Enzyme Kinetics

References

Koshland D. The Enzymes, v. I, ch. 7, Acad. Press, New York, 1959

Perutz M. Proc. Roy. Soc., B 167, 448, 1967

M.V. Volkenshtein, R.R. Dogonadze, A.K. Madumarov, Z.D. Urushadze, Yu.I. Kharkats. Theory of Enzyme Catalysis.- ''Molekuliarnaya Biologia'', Moscow, '''6''', 1972, pp. 431-439 (In Russian, English summary)

Cha, Y., Murray, C. J. & Klinman, J. P. Science 243, 1325-1330 (1989).

External links

http://us.expasy.org/enzyme/ links to Swiss-Prot sequence data, entries in other databases and to related literature searches

http://www.biochem.ucl.ac.uk/bsm/enzymes/links to the known 3-D structure data of enzymes in the Protein Data Bank

http://www.brenda.uni-koeln.de comprehensive compilation of information and literature references about all known enzymes; requires payment by commercial users

http://bioinformatics.weizmann.ac.il/cards/ extensive database of protein properties and their associated genes.

http://drnelson.utmem.edu/CytochromeP450.htmlsite lists over 4000 versions of enzymes from this cytochrome in plants and animals