Enzyme

An enzyme ( Ancient Greek coinage ἔνζυμον, énzymon ), formerly ferment (Latin fermentum ), is a substance that can catalyze one or more biochemical reactions. Almost all enzymes are proteins, the exception is the catalytically active RNA, such as snRNA. Their formation in the cell is therefore, as with other proteins via protein biosynthesis on the ribosome. Enzymes have important functions in the metabolism of organisms: You control the majority of biochemical reactions - from digestion through to transcription (RNA polymerase) and replication (DNA polymerase) to the genetic information.

  • 4.1 Energetic Foundations of Catalysis
  • 4.2 The active site - structural basis for catalysis and specificity
  • 4.3 Catalytic mechanisms
  • 5.1 Michaelis- Menten kinetics
  • 5.2 cooperativity and allostery
  • 5.3 Multi- substrate reactions
  • 5.4 enzyme inhibition
  • 6.1 Short-term adaptation
  • 6.2 Medium-term adaptation
  • 6.3 Long-term adaptation
  • 8.1 Enzymes in the art
  • 8.2 importance of enzymes in the medical diagnostic
  • 8.3 history of enzyme research

Word origin

Before 1878 they used to in the German language in the 15th century resulting from the Latin fermentum expression ferment. It means " fermentation medium " or " leaven" and was also used for the fermenter, fermentation and derived terms. 1833 of Eilhard Mitscherlich, the term ferment in connection with a substance which is not transformed at a reaction - but it is necessary to contact for a response - needed. 1878 led Wilhelm Friedrich Kühne today's neo-classical Greek art word enzyme ( ἔνζυμον, enzymon ) one derived from ἐν, en-, " in " and ζύμη, Zyme, which also means " the leaven " or " yeast ". This term then found its way into the international scientific community and is now part of the modern Greek language.

Nomenclature and classification according to IUPAC and IUBMB

Nomenclature

The IUPAC and the International Union of Biochemistry have together developed a nomenclature for enzymes, which classifies these homogeneous and numerous representatives of the group -containing molecules. To this end, developed the IUPAC principles of the nomenclature:

  • The enzyme name is to be explanatory, so the reaction that catalyzes the enzyme describe. ( For example, cholinesterase: An enzyme which hydrolyses the ester group in the choline molecule. )
  • The enzyme name should contain its classification ( see below). (Example: cholinesterase )

Further, a code system, the EC numbering system, developed in which the enzymes are classified under a code number of four figures. The first number refers to one of the six classes of enzymes. Lists all captured enzymes allow for faster retrieval of the specified enzyme codes, eg BRENDA. Although the codes are based on properties of the reaction, which catalyzes the enzyme, in practice, numerical codes, however, prove as unwieldy. More frequently used systematic, designed according to the rules above mentioned names. Problems of nomenclature arise, for example in enzymes that catalyze several reactions. For them, therefore sometimes there are several names. Some enzymes carry trivial names, do not let who realize that they are enzymes with said substance. As the name traditionally found wide use, they were partially retained ( examples: the digestive enzymes trypsin and pepsin of man).

Classification

Enzymes are classified according to the reaction they catalyze in six classes of enzymes:

  • EC 1: Oxidoreductases catalyze the redox reactions.
  • EC 2: transferases transferring the functional groups from one substrate to another.
  • EC 3: Hydrolases, the bonds using water column.
  • EC 4: lyases, catalyze the cleavage or synthesis of more complex products from simple substrates, but without consumption of ATP.
  • EC 5: isomerases that accelerate the conversion of chemical isomers.
  • EC 6: Ligases or synthetases that catalyze addition reactions using ATP. A reverse reaction ( fission) is usually energetically unfavorable and does not occur.

Some enzymes are able to catalyze a number of, sometimes very different reactions. If this is the case, they are attributed to several classes of enzymes.

Construction

Enzymes can be distinguished by their structure. While many enzymes consist of only one protein chain, known as monomers, there are other enzymes that oligomers of a plurality of subunits / protein chains. Some enzymes overlap with other enzymes to so-called multi-enzyme complexes together and co-operate or regulate one another. Conversely, there are also individual protein chains which can carry several different enzyme activities ( multifunctional enzymes ). Another possible classification as to its structure takes into account the presence of cofactors:

  • Pure protein enzymes consist exclusively of protein, the active site is formed only from amino acid residues and the peptide backbone. This group includes, for example, the digestive enzyme chymotrypsin and triose phosphate isomerase (TIM ) of glycolysis.
  • Holoenzymes ( altgr. ὅλος holos 'whole', 'complete' and enzyme ) consist of a protein moiety, the apoenzyme, as well as a cofactor, a low molecular weight molecule ( not a protein). Both together are important to the function of the enzyme. Organic molecules as cofactors are called coenzymes. If they are covalently bound to the apoenzyme, they are called prosthetic groups, otherwise even true co-substrate, since they are converted into equivalent amounts in the enzymatic reaction with the substrate. Co-substrates, for example adenosine triphosphate ( ATP) and nicotinamide adenine dinucleotide (NAD). ATP is often used as a source of energy for the reaction of protein kinases. NAD is used by enzymes, such as alcohol dehydrogenase, as an electron acceptor. Requires an enzyme metal ions (iron, zinc or copper ions), one speaks of a metalloenzyme. Lipoxygenase, for example, the iron and zinc containing carbonic anhydrase.

Function

Accelerate enzymes as biocatalysts biochemical reactions by lowering the activation energy that must be overcome, so that there is a material implementation. Theoretically, an enzymatic reaction is reversible, i.e., the products can be converted back into the starting materials. The starting materials ( reactants ) of an enzyme reaction, the substrates are bound in the so-called active site of the enzyme, it forms an enzyme -substrate complex. The enzyme then allows the conversion of the substrates in the reaction products are then released from the complex. Like all catalysts is the enzyme after the reaction front in the initial shape. Enzymes are characterized by high substrate and reaction specificity of, among numerous substances they only select the appropriate substrates and catalyze just one of many conceivable reactions.

Energetic basis of catalysis

Most biochemical reactions would take place without enzymes in living things only with negligible speed. The free reaction enthalpy () must be negative as with any spontaneously occurring reaction. The enzyme speeds up the chemical equilibrium - without changing it. The catalytic activity of an enzyme is based solely on its ability to lower the activation energy in a chemical reaction: this is the amount of energy that must be invested initially to set the reaction. While this is changing the substrate increasingly, it occupies an energetically unfavorable transition state. The activation energy is now the amount of energy that is required to force the substrate in the transition state. This is where the catalytic action of the enzyme: through non-covalent interactions with the transition state is stabilized this, so that less energy is required to bring the substrate in the transition state. The substrate can be converted into the reaction product much faster since it effectively a way " paved " is.

The active site - structural basis for catalysis and specificity

For the catalytic activity of an enzyme active site ( catalytic center ) is responsible. At this point, it binds the substrate, and thereafter "active" converted. The active site consists of folded parts of the polypeptide chain or reactive non-protein components ( cofactors, prosthetic groups ) of the enzyme molecule and requires a specificity of enzymatic catalysis. This specificity is based on the complementarity of the spatial structure and the surface potential interactions between enzyme and substrate. Leads to the formation of an enzyme -substrate complex.

The spatial structure of the active site causes only a structurally suitable substrate may be bound. Illustrative fits a specific substrate to the corresponding enzyme like a key into the appropriate lock (key - lock principle ). This is the reason for the high substrate specificity of enzymes. In addition to the lock and key model, the non-rigid Induced fit model exists: Since enzymes are flexible structures, the active site can be reshaped by interaction with the substrate.

Even small structural differences in spatial structure or charge distribution of the enzyme can lead to a substance similar to the substrate is no longer recognized as a substrate. Glucokinase example, but not by glucose as substrate, the related galactose. Enzymes may have different broad substrate specificity, so build alcohol dehydrogenases in addition to ethanol and other alcohols from Hexokinase IV and accepted in addition to glucose, other hexoses as a substrate.

The recognition and binding of the substrate is achieved through non-covalent interactions ( hydrogen bonding, electrostatic interaction or hydrophobic effects) between portions of the enzyme and the substrate. The binding of the enzyme must be strong enough to ( micromolar to millimolar concentrations) to bind the often low concentration substrate, but they must not be too strong, because the reaction does not end with the binding of the substrate. Important is an even stronger binding of the transition state of the reaction and thus its stability. Not infrequently, take two substrates participate in a reaction, the enzyme then must have the correct orientation of the reactants guarantee each other. These latter mechanistic peculiarities of an enzymatic reaction are the basis of the specificity of action of an enzyme. It always catalyzes only one of many conceivable reactions of the substrates.

Catalytic mechanisms

Although the mechanisms of enzymatic reactions in detail are multiform, use enzymes usually one or more of the following catalytic mechanisms.

Enzyme kinetics

The enzyme kinetics is concerned with the time course of enzymatic reactions. A key variable here is the reaction rate. It is a measure of the change in the substrate concentration in time, that is, for the amount of substance substrate, which is converted into a specific reaction volume per unit time (unit: mol / (L · s ) ). In addition to the reaction conditions such as temperature, salt concentration and pH of the solution, it depends on the concentration of the enzyme, the substrates and products, and effectors (activators or inhibitors).

In connection with the reaction rate is the enzyme activity. It indicates how much active enzyme is in an enzyme preparation. Units of enzyme activity are unit (U) and katal ( kat ), wherein 1 U is defined as that amount of enzyme which converts one micromole of substrate per minute under specified conditions: 1 U = 1 mol / min. Katal is rarely used, however, is the SI unit of enzyme activity: 1 kat = 1 mol / s Another important parameter for enzymes is the specific activity ( activity per unit mass, U / mg). That shows how much of the total protein in the solution is really the desired enzyme.

The enzyme activity measured is proportional to the reaction rate and thus strongly depends on the reaction conditions. It increases with the temperature according to the RGT control: increasing the temperature by about 5-10 ° C resulting in a doubling of the reaction rate and thus the activity. However, this applies only for a limited temperature range. At an optimal temperature is exceeded, there is a steep drop in the activity by denaturation of the enzyme. Changes in pH of the solution often has a dramatic effect on the enzyme activity, because it can affect the charge of individual amino acids important for catalysis in the enzyme. Beyond the optimum pH decreased enzyme activity and eventually comes to a halt. The same applies to the salt concentration or the ionic strength in the area.

Michaelis- Menten kinetics

A model for the kinetic description of simple enzyme reactions is the Michaelis- Menten kinetics (MM- theory). It provides a link between the reaction velocity v of an enzyme reaction and the enzyme and substrate concentration [ E0 ] and [ S]. It is based on the assumption that an enzyme with a substrate molecule forms an enzyme- substrate complex and this decays in either enzyme and product or to its original constituents. What happens quickly, depending on the respective velocity constants k.

The model predicts that, with increasing substrate concentration increases the reaction rate. This is done at first linearly and then levels off until a further increase in the substrate concentration has no effect on the rate of the enzyme, since this is already working at maximum speed Vmax. The MM equation is as follows:

The parameters Km ( Michaelis constant) and kcat ( turnover number ) are suitable enzymes to characterize kinetically, that is, to make statements about their catalytic efficiency. Is Km example, very low, it means the enzyme reached at low substrate concentration exceeds its maximum speed and thus is very efficient. At low substrate concentrations, the specificity constant kcat / Km is a more appropriate measure of the catalytic efficiency. It reaches values ​​of more than 108 to 109 M-1 s-1, the reaction rate is only limited by the diffusion of substrate and enzyme molecules. Every random contact of enzyme and substrate leads to a reaction. Enzymes that achieve such efficiency, called " catalytically perfect".

Cooperativity and allostery

Some enzymes do not show the hyperbolic saturation curve as predicted by the Michaelis- Menten kinetics, but a sigmoidal saturation behavior. Something was first described with binding proteins such as hemoglobin, and is interpreted as a positive cooperativity multiple binding sites: the binding of a ligand (substrate molecule) affects more binding sites in the same enzyme (often but in other sub- units) in their affinity. In case of positive cooperativity has a binding protein with many free binding sites a weaker affinity than a mostly occupied protein. Binds the same ligand binding to all centers, one speaks of a homotropic effect. The cooperativity is closely linked with the allostery in enzymes. Under Allosterie mean the presence of additional binding sites ( allosteric centers) in an enzyme, apart from the active site. Binding effectors (not substrate molecules ) to allosteric centers, is a heterotropic effect is present. The allostery, although conceptually distinct from the cooperativity, yet they often occur together.

More substrate reactions

The previous considerations are valid only for reactions in which a substrate is converted to a product. However, many enzymes catalyze the reaction of two or more substrates or co-substrates. Likewise, several products can be formed. For reversible reactions, the distinction between substrate and product is already relatively. The Michaelis- Menten theory is valid for one of a plurality of substrates, when the substrates with the other enzyme is saturated.

For multi- substrate reactions, the following mechanisms are conceivable:

  • Random mechanism (English random): The sequence of the substrate binding is random.
  • Parent mechanism ( engl. ordered ): The order of binding is established.

Enzyme inhibition

The enzyme inhibition ( inhibition) is defined as the reduction in the catalytic activity of an enzyme by a specific inhibitor (inhibitor ). Is fundamentally different to the irreversible inhibition, wherein an inhibitor undergoes a non-reversible under physiological conditions associated with the enzyme (such as penicillin with D-alanine transpeptidase ) of the reversible inhibition, in which the formed enzyme-inhibitor complex again can decay into its constituents. The reversible inhibition, a distinction again between

  • Competitive inhibition - the substrate competes with the inhibitor for binding to the active site of the enzyme. But the inhibitor is not enzymatically convertible, thereby stopping the enzyme work by blocking the active site;
  • Allosteric ( non- competitive inhibition ), - the inhibitor binds to the allosteric center, thereby changing the conformation of the active site, so that the substrate can no longer bind therein;
  • Uncompetitive inhibition - of the inhibitor binds to the enzyme -substrate complex, thereby preventing the catalytic conversion of substrate to product.

Regulation and control of the enzyme activity in the organism

Enzymes act together in a complex network of metabolic pathways in living organisms. In order to adapt to fluctuating internal and external conditions optimally, a fine regulation and control of metabolism and the underlying enzymes is necessary. Under regulation refers to processes that serve to maintain stable internal conditions under changing environmental conditions ( homeostasis). As a control is referred to changes that take place due to external signals (eg, by hormones). There are fast / short, medium and slow / long -term regulation and control processes in the metabolism:

Short-term adaptation

Rapid changes in the enzyme activity take place as a direct response of the enzymes to altered levels of metabolites, such as substrates, products or effectors (activators and inhibitors ). Enzyme reactions are close to equilibrium, are sensitive to changes in the substrate and product concentrations. Accumulation of substrate accelerates the forward reaction, accumulation of product inhibits the forward reaction and promotes the reverse reaction (competitive product inhibition). But in general a greater role in the metabolic regulation and control is attributed to the irreversible enzyme reactions.

Of great importance is the allosteric modulation. Substrate or effector molecules that occur in metabolism, bind to allosteric centers of the enzyme and alter its catalytic activity. Allosteric enzymes are composed of several subunits (either from the same or from different protein molecules). The binding of substrate or inhibitor molecules to a subunit resulting in conformational changes in the total enzyme, which alter the affinity of the remaining binding sites on the substrate. An end-product inhibition ( feedback inhibition ) occurs when the product of a reaction chain on the enzyme at the beginning of this chain acts allosterically inhibiting. This automatically creates a loop.

Medium-term adaptation

A common form of metabolic control is the covalent modification of enzymes, particularly phosphorylation. As if by a molecular switch, the enzyme may, for example, according to a hormonal signal can be switched on or off by phosphate - transferring enzymes ( kinases). The introduction of a negatively charged phosphate group pulls structural changes in the enzyme by itself and can in principle promote active and inactive conformations. The cleavage of the phosphate group by phosphatases reverses this process, so that a flexible adaptation of the metabolism to changing physiological requirements is possible.

Long-term adaptation

As a long -term response to changing demands on the metabolic enzymes are selectively degraded or newly formed. The formation of new enzymes is controlled by the expression of its genes. Such a type of genetic regulation in bacteria describes the operon model of Jacob and Monod. The controlled degradation of enzymes in eukaryotic cells may be accomplished by ubiquitination. The attachment of polyubiquitin chains to enzymes that catalyzed by specific ubiquitin ligases, this marked for degradation in the proteasome, a " garbage disposal " of the cell.

Biological Significance

Enzymes are not to be underestimated biological meaning, they play a central role in the metabolism of all living organisms. Virtually every biochemical reaction is accomplished and controlled by enzymes. Well-known examples are glycolysis and the citric acid cycle, respiratory chain, photosynthesis, transcription and translation, and DNA replication. Enzymes not only act as catalysts, they are also important regulatory and checkpoints in the metabolism.

The importance of enzymes is not limited to the metabolism, even when the stimulus recording and sharing are important. In signal transduction, which is the placement of information within a cell receptors are frequently involved with enzymatic function. Even kinases such as tyrosine kinases and phosphatases play a crucial role in the transmission of signals. The activation and deactivation of the information carrier, that is the hormone done by enzymes.

In addition, enzymes involved in the defense of one's own organism, for example, are a variety of enzymes such as serine proteases of the complement part of the innate immune system of humans.

Error in enzymes can have fatal consequences. Such enzyme defects, the activity of an enzyme is reduced or no longer exist. Some defects are genetically inherited enzyme, i.e., the gene that encodes the amino acid sequence of the corresponding enzyme, comprises one or more mutations or absent. Examples of hereditary enzyme defects are phenylketonuria and galactosemia.

History, use and occurrence in everyday life

Enzymes are valuable tools of biotechnology. Their applications range from the production of cheese ( rennet ) on the enzymology through to genetic engineering. For certain applications, scientists are now developing targeted powerful enzymes by protein engineering. In addition, one designed a novel form of catalytically active proteins, catalytic antibodies which have been mentioned, due to their similarity to enzymes abzymes. Also ribonucleic acids ( RNA) can be catalytically active; These are then referred to as ribozymes.

Enzymes are needed, among other things in the industry. Detergents adds to lipases ( lipolytic enzymes), proteases ( proteolytic enzymes) and amylases ( amylolytic enzymes) added to increase the cleaning power, since these enzymes decompose the corresponding spots. Enzymes are also used in the preparation of some drugs, and insect repellent. In cheese making, rennet which interacts with, an enzyme obtained from calf stomachs. Many enzymes can be produced with the help of genetically modified microorganisms today.

The enzymes contained in raw pineapple, kiwifruit and papaya prevent the solidification of cake gelatin, an undesirable effect if, for example, a fruit cake containing raw pieces of this fruit, to be coated with a solid cake gelatin coating. The soft Remain About the casting does not occur with the use of fruit from cans, they are pasteurized, the protein degrading enzymes are inactivated.

When peeling of fruits and vegetables plant cells are injured and released enzymes in the sequence. This allows the peeled Good ( well seen in apples and avocados ) are brown by enzymatically assisted reaction of flavonoids or other sensitive ingredients with atmospheric oxygen. The addition of lemon juice acts as an antidote. The ascorbic acid present in the lemon juice prevents oxidation or reduced already oxidized compounds ( the addition of ascorbic acid as a food additive ).

In medicine, enzymes play an important role. Many drugs inhibit enzymes or enhance their impact in order to cure a disease. The most prominent representative of such drugs is probably the acetylsalicylic acid, which inhibits the enzyme cyclooxygenase and thus relieves pain among others.

Enzymes in the art

The following table gives an overview of the uses of enzymes. To produce protein # see production of specific proteins.

Importance of enzymes in the medical diagnostic

The diagnostic uses enzymes to detect diseases. In the test strip is for diabetics, for example, an enzyme system which produces a fabric under the influence of blood sugar, the content thereof can be measured. Thus, the blood sugar level is measured indirectly. We call this approach a " enzymatic measurement". It is also used in medical laboratories for the determination of glucose (blood glucose ) or alcohol. Enzymatic measurements are relatively simple and inexpensive to use. It makes use of the substrate specificity of enzymes advantage. Thus, the body fluid to be analyzed is added an enzyme that may convert the substrate to be measured specifically. To the resulting amount of the reaction products then can be read how much of the substrate in the body fluid was present.

In human blood, a number of enzymes based on their activity are directly measurable. Circulating in the blood enzymes originate partly specific organs. It can therefore be drawn on the basis of lowering or increasing of enzyme activities in the blood conclusions to damage certain organs. Thus, an inflammation of the pancreas can be recognized by the greatly increased activity of the lipase and pancreatic amylase in the blood.

History of enzyme research

The scientific study of enzymes began in 1833 when the French chemist Anselme Payen diastase the first enzyme discovered at all. Another milestone set the studies on the enzyme specificity of Emil Fischer dar. He postulated that enzymes and their substrate behave like a lock and matching key. 1897 Eduard Buchner discovered on the basis of alcoholic fermentation, enzymes that can also act as a catalyst without the living cell. Beginning of the 20th century was done very much in enzyme research. The most important scientists of this period was the German chemist Otto Röhm. He isolated the first enzyme and procedures developed for the enzymatic tanning leather, fruit cleaning and a range of diagnostic applications. Leonor Michaelis and Maud Menten pioneered in the study of enzyme kinetics.

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