Enzymes are Proteins: A Definitive Guide of 4000+ Words (Updated)
Enzymes are Proteins – The Guide Encompasses Introduction, History, Structure, Active Site, Enzyme Action, Functions, Models, Factors, Inhibitors, Classification, Nomenclature
Enzymes are proteins that speed up the reactions by acting as catalysts. They increase the rate of reaction without being consumed or permanently altered by themselves. In Greek enzymes are divided into sections “En” which means “in” and second part is “zyme” means living. The study of enzymes is known as enzymology. (Enzyme Definition)
Introduction to Enzymes
Physical nature of life is based on the making and breaking of different composed at all levels involved in the metabolism. Thousands of compounds must be shaped to produce the organelles and other structures present in living organisms. The formation may involve the making of large molecules from smaller molecules or vice versa, moreover all these reactions should take place at moderate temperature and with a speed good enough to support life. It is a common laboratory observation that reactions taking place within a cell if accomplished outside the cell needs very high temperature and there is no control on the speed of the reaction. Why the same reactions need different environments to accomplish when taking place inside the cell. What is the missing connection between living and non-living environment? Initially people thought that it is because of the life itself. A living thing can do what cannot be done by the non-living conditions.
History of Enzymes
The enzymes were first discovered by a French chemist named Anselme Payen. He named them as diastase in 1833. A few decades later, Louis Pasteur while studying the fermentation of sugar to alcohol by yeast discovered that there is an essential force, he called it ferments that only function within living organisms. The term enzyme was first used in 1878 by Wilhelm Kuhne, a German physiologist. Thus, after lot of hard work the scientists were able to find the missing connection. They were able to demonstrate that it is not the life but a large biomolecule made of amino acids, which behaves as a catalyst in the functioning of the cell. These large molecules not only speed up the reactions but also make them happen at moderate temperature which can support life. This biomolecule takes part in all the formations and breakdown of compounds which follow specific pathways.
All the reactions require the biomolecule as definite controlling system. The control should be for the direction, speed and specificity of the undergoing reaction. Cells control all operations by producing the proper biomolecuse, named Enzymes. The enzymes are proteins that always produced in the proper amounts, at the proper time and the proper place where they are needed and according to research their are different factors that affect enzyme activity i.e., pH, Temperature, Enzyme and Substrate Concentration etc.
As mentioned above in the enzymes definition, they are required by the biomolecules, because nearly all chemical reactions of life are so slow without catalysts that they cannot sustain life. The enzymes generally speed up reaction rates by factors between 108 and 1020 and thus made the life process possible.
Structure of Enzymes
One of the basic qualities of enzymes is that they are specific in action. If we elaborate the specificity, we will understand that specificity is because of their power of recognizing the substrate from a sea of molecules. How they do this. They do this because of the specific structure of the substrate. This means that enzymes are structure specific in nature. This structure specificity of the enzymes is based on their own specific structure when proper substrate colloids with a proper enzyme having proper structure, they fit in to carry out the phenomena of catalysis. Thus, for the proper understanding of Enzymes and enzyme action we need to understand their structure and chemical nature.
Enzymes are proteins, which are giant and of special type, that are composed of hundreds of amino acids. These amino acids are joined together in a linear fashion to produce chains. The union of amino acids into polypeptide chains of proteins occurs through peptide bonds involving the carboxyl group of one amino acid and the amino group of the next amino acid. These chains coil upon themselves to form globular three-dimensional structure. These structures may be the result of a single chain of amino acids or more than one chain may give rise to these structures. The polypeptide chains may be identical thus known as homopolymers (homo-same; poly-many; mers-parts). Mostly the chains in enzymes are different. In such case they are known as heteropolymers (hetero-different; poly-many; mers- parts).
About 90 % of the Co factor enzymes are pure protein i.e. composed of amino acid chains only. In the rest of 10% the enzymes protein is the major part associated with a small non-proteineous part. In such enzymes the protein parts of enzymes are known as Apoenzyme, and a much smaller, organic non protein portion called co factor.
Apoenzyme and Co factor: Apoenzyme and co factor joins together to form functional enzyme which is known as Holoenzyme. Co factor is essential for the catalytic activity of the holoenzyme.
Co-factor can be further divided on the basis of their attachment with apoenzyme into three categories.
- a) A co-factor is called prosthetic group when it is tightly attached to the apoenzyme by covalent bonds.
- b) Sometime the co factor is loosely attached with the apoenzyme through weak forces. In this status the non proteineous part is called coenzymes.
The co-factor may still be a metallic ion such as magnesium, and potassium for their activity. The metal ions are often called metal activators. For example, Hexokinase requires Mg++ ions.
Active Site of Enzyme
Most of the enzymes are composed of about 250 amino acids. As the attachment of amino acids grows the peptide chain length increases. Dipeptide changes to tripeptide, tripeptide into tetrapeptide and so on. Every added amino acid although increases the chain length but this does not mean that it gives rise to a straight structure. Instead the chain folds and bends upon itself to give rise to a globular three-dimensional structure. The resulting three-dimensional structure of enzyme which is a functional unit always have a depression or groove on its surface which is known as active site of an enzyme in previous above diagram.
This active site is mostly composed of about 10 to 12 amino acids brought close to each other by bending and folding of the polypeptide chain. Rest of the amino acids plays a part in the stabilizing the enzyme structure even during enzyme action. Active site engages the substrate for enzyme action during enzyme catalysis.
Types of Active Site
The active site is further divided into types:
- Binding site
- Catalytic site
Binding site recognizes the substrate and helps in the binding of the substrate/inhibitor while catalytic site when engages does the catalysis.
Enzyme action mechanism involves making or breaking of biomolecules. The enzyme’s role in the reaction is of biocatalyst. All the reactions, which can otherwise take place if provided with enzyme, simply increase their speed. Enzymes can speed up the reaction rate but they cannot force a reaction to take place. The mechanism involved in the reaction remains the same enzymes only speed up the process. How it is possible? To understand this, we have to first understand how reaction takes place and then how enzyme speed up the process.
The process of making and breaking (reaction) depends upon the change in energy level. In a chemical reaction usually the most energetic molecules are able to undergo chemical reactions. Such molecules become more energetic than others of the same kind by being subjected to different numbers and types of collisions. A rise, in temperature increases the possible collision rate thus reaction rate is increased.
Energy of Activation
Molecules that have enough energy undergo reaction are converted to products. The energy required for the molecules to undergo a change in known as energy of activation. In laboratory experiments energy is provide to the system to increase the activation energy of the molecules to produce products. But in the living system it is not possible to increase the temperature to such elevated state of energy, which could not support life.
The only possibility to produce products without increasing the energy of the molecule is to decrease the activation energy of the system. This is possible only because of the mediation of enzyme. It is through enzymes that life carries out such reactions at relatively low temperature suitable for life. The enzymes being proteins do this job by lowering the energy of activation so that a much greater fraction of the substrate molecules has sufficient energy to react without a temperature increase, thus substrates are converted into products. A careful study of graph can further help in the understanding of this enzyme action mechanism.
Enzyme Substrate Complex
For an enzyme to decrease the energy of activation during the course of the reaction, the enzyme must combine temporarily with the, substrate or substrates to form an enzyme substrate complex.
E + S → ES Complex → E + P
E = Enzyme
S = Substrate
ES Complex = Enzyme Substrate Complex
P = Products
The formation of enzyme substrate complex plays the key role in the reduction of activation energy of the reaction. The enzyme substrate complex thus formed decreases the activation energy by changing the status of substrate. At this point we should not forget that substrate binds with enzyme only at active site, which is not a flat surface but a grove. Once substrate combines to form enzyme substrate complex. The resulting structure is neither enzyme nor substrate but a complex.
In the groove micro-environment is modified to the extent that forces stabilizing the molecules (substrate and enzyme) within this complex are modified. Thus, effects the forces of the substrate, causing some of the substrate bonds to break; then the bonds rearrange to form stable structure the products, much more rapidly than in the absence of the enzyme. Enzyme molecule remains stable only because of its size, which is about hundred times larger than substrate.
Bonding involved in the formation of ES Complex
The kinds of bonds between enzymes and substrates can be covalent, ionic, hydrogen, and Vander Walls. The covalent and ionic bonds are most important with respect to the energy of activation for a reaction, but the more numerous hydrogen bonds and Vander Walls interactions contribute to the structural orientation of the enzyme substrate complex. Even when strong covalent bonds are formed, they are usually very rapidly broken to release new product molecules. Both covalent and non-covalent bonds between enzyme and substrate are formed between parts of the R portions of amino acid residues in the enzyme, not between the carbon and nitrogen atoms involved in the peptide bonds. Thus, enzymes must have the proper amino acids (composition) and must have them in the right places (sequence) for its proper functioning.
Function of Enzymes
The points and diagram below will simplify the function of enzyme.
- They speed up reactions and act as catalysts. As they make reactions millions times faster.
- Help in digestion, Break large complex molecules into smaller molecules i.e., glucose which is utilized by organisms body.
- Helps in unwinding the DNA and copying of it inside the cell.
Model of Enzyme Action
Lock and Key Model – Emil Fischer
Emil Fischer proposed lock and key model in about 1884. The basic idea of the model lies in the specific nature of the enzyme action. As only a specific key can open a specific lock. Fisher thought that enzyme being specific in nature must be behaving like a lock and key.
According to this model of enzyme action, the substrate and enzyme forms a complex before substrate’s conversion to products. Just as one specific key open a specific lock. This idea gives a rigid lock and key union between the enzyme and substrate. The complex is established at active site.
According to this model, the shape of the active site is not modified before, during or after the reaction, and the enzyme is given off as such after the termination of the reaction.
Objection on Lock and Key Model
We understand that biomolecules, enzyme and substrate cannot behave like rigid structures at the time of complex formation as proposed in this model. If the active site were rigid and specific for a given substrate reversibility of the reaction would not occur because the structure of the product is different from that of the substrate and would not fit the enzyme well, as contrasted to a rigidly arranged active site.
Induced Fit Model – Daniel E. Koshland
Although basic idea remains the same, new data added to the understanding of enzyme action forced a new model known as Induced fit model or hand and glove model.
Daniel E. Koshland, Jr. (1973) found evidence that the active site of enzymes can be induced by close approach of the substrate or substrates (or product or products, when the reaction reverses) to undergo a change in conformation (shape) that allows a better combination of the substrate with the enzyme. This idea is now widely known as the induced fit model/hypothesis, or hand and glove model. It is explained through pictures for an enzyme with only one substrate. If two or more substrates are involved, the principle remains the same, with each substrate contributing.
The basic idea captured in this model explains. that as glove has its own shape but when it is put on its shape is modified with respect to the fingers moving in place inside the glove. In the same fashion the shape of the substrate and enzyme are modified to some extent to allow for a better fit in position between substrate and enzyme to facilitate the reaction. Apparently ‘the structure of the substrate is also changed during many cases of induced fit, allowing a more functional enzyme substrate complex.
Factors Affecting Enzymes Activity
Factors Affecting Enzymes Activity: Catalysis occurs only if enzyme and substrate form a transient complex i.e. enzyme substrate complex. Whereas reaction rate depends on the number of successful collisions between them, which in turn depends upon their concentrations.
Taking into consideration these rules we will try to understand the influence of different factors affecting enzymes activity and how these factors can affect the enzyme action.
One enzyme molecule can transform one molecule bf its substrate per unit time. Thus, the rate of reaction depends directly on the amount of enzyme. If enzyme is increased to two folds the reaction rate will be doubled if enough substrate is present.
The number of active sites increases by the increase of the enzyme molecules. So, more active sites will convert more substrate into products in the given period of time. Thus, an increase in the reaction rate is directly proportional to the amount of enzyme present (Fig.5.1)
Substrate concentration has two-fold effect on the reaction rate. At low concentration of substrates, the reaction rate is directly proportional to the substrate available. If the enzyme is kept constant and the amount of substrate is increased, a point is reached when a further increase in the substrate does not increase the rate of reaction any more. This is because at high substrate level all the active sites of the enzyme are occupied and further increase in the substrate does not increase the reaction rate (Fig.5.4).
Effect of pH on Enzyme Activity
Optimum pH refers to the narrow range of pH on which every enzyme function most effectively. Since enzymes are proteins slight change on either side of the optimum pH modifies its effect on the substrate. Change in pH has two ways effect on the reaction rate. A slight change in pH can stimulate ionization of the amino acid’s residences at the active center which may prevent its proper attachment with the substrate. Same is true for the substrate. Moreover, it also affects the ionization of the substrate. Under these changed conditions enzyme activity is either restricted or blocked altogether. This reduces the reaction rate. Extreme change in pH can cause the bonds in the enzymes to break resulting in the denaturation of the enzyme. This checks the reaction rate forever (Fig.5.3).
Effect of Temperature on Enzyme Activity
As opposed to mammals and birds, plants cannot regulate their temperatures. As a result, all reactions in them are strongly influenced by external temperatures. In general, reactions catalyzed by enzymes increase with temperature from 0°C to 35 or 40°C (Fig.5.10). The increase in reaction rate is partly because heat increases the number of molecules that have energy equal to or greater than the energy of activation. Yet because the reaction rate so strongly depends on catalysis by the enzyme, temperature also affects reactions by changing the shape of the enzyme. The enzyme’s shape determines its ability both to combine with the substrate and to cause catalysis thus effects directly on reaction rate.
An organism’s growth and reproduction vary greatly with temperature, and for any given species this may depend on the optimum temperature for action of certain enzymes that control rate-limiting reactions.
If the temperature is raised above a certain value, the reaction rate begins to decrease because of denaturation of most enzymes (Fig.5.2). Similarly, low temperatures can also denature certain enzymes. Various enzymes, even from a single species, often differ greatly in their response to temperature. This means that at any given temperature some enzymes function optimally or nearly so, whereas many others function far less than their optimal status. This phenomenon appears to be especially important in species that are sensitive to chilling. This difference in the temperature effect on different enzymes helps in determine the environment in which different species can live. For example, the overall temperature optimum for the process of photosynthesis (catalyzed by many enzymes) in alpine and arctic plants is often 10 to 15°C, whereas the optimum for maize plants is near 30°C as given in below chart.
The rate of an enzymes reaction can be determined by measuring the rate of disappearance of substrate or the rate of product appearance, or both. By either method the reaction is usually observed to proceed more slowly as time passes. This decrease in rate is sometimes caused by denaturation of the enzyme while the reaction is being measured, but other factors are also involved. One of the most important factors is the continuous decrease in concentration of substrate and/or accumulation of products. As products accumulate, their concentrations sometimes become high enough to cause appreciable reversibility of the reaction, provided that the relative chemical potentials of products and reactants allow reversibility. In some cases, i.e., in forward reaction the reaction products inhibit catalysis in enzymes by combining with the enzyme in such a way that further formation of the enzyme substrate complex is inhibited.
Enzyme Inhibitors Definition: Those chemical substances which can react in the place of substrate with an enzyme but are not converted into products and blocks the active site temporarily or permanently are called inhibitors. From inhibitors, comes the process of enzyme inhibition.
Enzyme Inhibition Definition: Inhibition is the capacity of the inhibitors to retard or stop the enzyme catalysis. Substrate for example poisons, anti-metabolites, antibiotics and some drugs act as inhibitors for certain enzymes and thus stop their activity.
Their are two types of inhibitors.
- Irreversible inhibitor
- Reversible inhibitors
This type of inhibitor check the reaction rate by occupying the active site or destroying the enzyme’s globular structure. They occupy the active site directly by having a covalent bonding to any of the amino acids situated in the active site or they may physically block the active site.
The irreversible inhibitor may also combine with sulphydral (-SH) groups and destroy the disulphide bridges, which are essential for maintaining the globular structure of the enzyme. As a result, enzyme molecule is denatured.
This is the chemical, which react with the active site temporarily. They form weak interaction and either retard or stop the enzyme activation. Effect of reversible inhibitors can be neutralized completely or partly by an increase in the concentration of the substrate.
Four major types of reversible inhibitors are known to exist. They are:
- Non Competitive
- End product inhibition
a) Competitive Inhibitors: Because of their structural similarity with substrate they are associated with enzymes at active site. They are able to attach itself at binding site but are not able to activate its catalytic site, thus given off. The degree of inhibition depends upon the relative concentration of substrate and inhibitor. By increasing substrate concentration reaction rate can be increased, thus the effect of inhibitor can be reduced to almost zero. A very good example is Malonic acid and Succinic acid, which compete for same site (Fig.5.11).
b) Non Competitive Inhibitor: These types of inhibitors do not compete with the substrate for the attachment on the active site. Their function is to form the enzyme inhibitor complex at a point other than the active site. They alter the structure of the enzyme in such a way that even if genuine substrate tries to bind the active site, catalysis fails to take place.
As substrate and inhibitor are not competing for the same site so an increase in substrate concentration has no effect on the reaction rate.
c) Uncompetitive Inhibitor: They always combine with the Enzyme Substrate Complex and not with the enzyme when enzyme is alone. As a result, the reaction rate is inhibited. This inhibition is also not affected with an increase in the substrate concentration.
d) End Product Inhibition (Negative Feedback Inhibition): This inhibition is caused by the interaction of a product with an enzyme in the sequence of its formation. In feedback inhibition the inhibited enzymes may very often be an allosteric enzyme (allo means “other”; that is, different from the active site). This type of inhibition is mostly of competitive type (see below diagram).
Allosteric Enzymes and Feedback Control
We have mentioned that numerous foreign ions or molecules (inhibitors) can inhibit enzymatic action, in most cases by altering the configuration of the enzyme so that it cannot effectively form a complex with the substrate. However normal cellular constituents can also alter several enzymes thus resulting in the decreases or increases in their functions. Such effects are important mechanisms for homeostatic control at the metabolic level. They help organisms produce only the proper amount of the compounds they need. The most common case is inhibition of a particular reaction by a metabolic that is chemically unrelated to the substrate with which the enzyme reacts.
To understand this, consider an example in which a compound A is converted by series of enzymatic reactions via intermediates B, C, D, and E to an essential product F. After this number of reactions, compound F no longer bears much structural resemblance to A. Nevertheless, F can sometimes reversibly combine with the first enzyme to inhibit its combination with A. This is an example of feedback inhibition or end product inhibition. Its advantage is that it provides a rapid and sensitive mechanism to prevent over synthesis of compound F. Because the feedback inhibition occurs only after F has built up to a level that is sufficient for cellular needs. Later, when the amount of F in the cell has been reduced (by incorporation into a structural component of the cell), F molecules dissociate from enzyme number 1 and allow it to become active again. Cases of feedback inhibition nearly always involve action of a product of a metabolic pathway upon the first enzyme of that pathway.
A well-studied example of feedback inhibition in plants occurs in formation of the nucleotide uridine monophosphate (UMP), which begins with aspartic acid and carbamyl phosphate. The pathway requires five enzyme-catalyzed steps, but only the first enzyme, aspartic transcarbamylase, is susceptible to feedback control by UMP; no other reaction is blocked in a similar way by reactants and products in the pathway.
Enzymes that combine with and respond (either negatively or positively) to small molecules such as F are called allosteric enzymes. The sites at which combination with the smaller molecules occurs are called allosteric sites.
Classification of Enzymes
Classification of Enzymes: More than 5,000 different enzymes have been discovered in living organisms, and the number grows as research continues. The most important properties of enzymes, so far studied are their specificity. Each enzyme acts on a single substrate (reactant) or a small group of closely related substrates that have virtually identical functional groups that are capable of reacting.
Nomenclature of Enzymes
In the beginning when the data regarding the enzymes was very limited each enzyme was given a trivial name. For example, pepsin, trypsin gastrin etc. As the data increased a need for some rule was badly felt. Thus, all enzymes were named according to a standardized system.
The Suffix Ase
All enzymes were named to end in the suffix ‘ase’. These names characterize the substrate or substrates acted upon and the type of reaction catalyzed. For example, cytochromeoxidase, an important respiratory enzyme, oxidizes (removes an electron from) a cytochrome molecule. Malic acid dehydrogenase removes two hydrogen atoms from (dehydrogenates) malic acid. These common names, although conveniently short, do not give sufficient information about the reaction catalyzed. For example, neither identifies the acceptor of the removed electron or hydrogen atoms.
The International Union of Biochemistry 1961 lists longer but more descriptive standardized names for all well-characterized enzymes. As an example, cytochrome oxidase is named cytochrome c: O2 oxidoreductase, indicating that the particular cytochrome from which electrons are removed is the c type and that oxygen molecules are the electron acceptors. Malic acid dehydrogenase is -called L malate:NAD oxidoreductase, indicating that the enzyme is specific for the ionized L form of malic acid (malate) and that a molecule abbreviated as NAD is the hydrogen-atom acceptor.
Classes of Enzymes
A systematic approach with new names was first suggested in 1965 and revised n 1972. According to this system all enzymes are grouped in six main classes. The main classes of enzymes and the type of chemistry they participate is given below.
The system is designed to zero on the specific identity of each enzyme by dividing each main class into subclasses and sub-subclasses. By using a numbering system throughout the scheme, each enzyme can be assigned a numerical code, such as 18.104.22.168, where the first number specific subclass and sub-subclass respectively and the final number represents the serial listing of the enzyme in its subclasses. For example, histidine decarboxylase (traditional name) is identified as histidine decarboxy-Iyase, 22.214.171.124; alcohol dehydrogenase as alcohol:NAD oxidireductase, 126.96.36.199; urease as urea amidohydrolase, 188.8.131.52.
Let us see how it works,
Lyases (cleavage ) Class
4.1. Carbon-carbon Iyases (cleavage of C-C bonds) Subclass
- 4.1.1. Carboxyl-Iyases (cleavage of C-COO– bond) Subclass
- 184.108.40.206. histidine carboxylase (cleavage of C-COO bond in histidine) name
- 4.1.2. Aldehyde-Iyases
4.2. Carbon-oxygen lyases (cleavage of C-O bond)
4.3. Carbon nitrogen Iyases (cleavage of C-N bond)
4.4. Carbon-Sulphur Iyases (cleavage of C-S bond)
4.5. Carbon halogen Iyases
4.6. Phosphorus-oxygen Iyases
The use of this modern classification of enzymes is required in most of the professional research journals. However, the older system is still widely practiced, mostly in monographs and textbooks.