6. BIOMOLECULES�Part - 3

-Created By-

Shri. Deshmukh A. B.

Asst. Teacher, Biology

Agasti Art’s, Commerce and Dadasaheb Rupwate

Science College, Akole

E. Enzymes :

• Thousands of different chemical reactions take place automatically at a given time in a tiny living cell.
• The reactions take place at the body temperature.
• If these enzymes were not present in the cell,
• either the reactions would not occur or
• if they occur they would occur at a very very slow rate.

Know the scientists

• German chemist Edward Buchner discovered enzymes by accident.
• Buchner discovered that living cells were not necessary but that yeast extract could bring about fermentation.
• He then coined the term Enzyme (Gk. En = in, zyma = yeast i.e. In yeast).
• This term is now commonly used for all biocatalysts.

• Each enzyme catalyzes a small number of reactions, specifically perhaps only one.
• e.g. Urea + Water Ammonia + CO₂
• The substance upon which an enzyme acts is termed as the substrate.
• The enzymes which act within the cell in which they are synthesized are known as endo -enzymes
• e.g., enzymes produced in the chloroplast and mitochondria,
• If they act outside the cell in which they are synthesized, they are known as exo-enzymes.
• e.g., enzymes released by many fungi.
• These enzymes, synthesized by living cell, retain their catalytic property even when extracted from cells.

Urease

Do you know ?

• Rennet tablets used for coagulating milk protein casein (cheese) contain renin enzyme that is obtained from the stomach of calf.

Nature of Enzymes :

• On the basis of chemical composition, enzymes can be put into two categories.
• (i) Purely proteinaceous enzymes :
• e.g. proteases that spilt protein

• (ii) Conjugated enzymes :
• These are made up of a protein to which a non-protein prosthetic group is attached.
• The prosthetic group is firmly bound to the protein component by chemical bonds and is not removed by hydrolysis.
• If the prosthetic group is removed the protein part of the enzyme becomes inactive.

• There are enzymes which require certain organic compounds and inorganic ions for their activity.
• The organic compounds that are tightly attached to the protein part are called co-enzymes whereas the inorganic ions which are loosely attached to the protein part are called co-factors.
• Some of the organic co-enzymes are
• nicotinamide adenine dinucleotide (NAD) and
• flavin mononucleotide (FMN).

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• Inorganic ions of metals which act as co-factors include
• Magnesium
• Copper
• Zinc
• Iron - (Fe++) is a co-factor of enzyme catalase
• Manganese - is a co-factor of peptidases
• Often metal co-factors are referred to as enzyme activators.

Properties of Enzymes :

• Proteinaceous Nature :
• All enzymes are basically made up of protein.
• Three-Dimensional conformation :
• All enzymes have specific 3-dimensional conformation.
• They have one or more active sites to which substrate (reactant) combines.
• The points of active site where the substrate joins with the enzyme is called substrate binding site.

• Catalytic Property :
• Enzymes are like inorganic catalysts and influence the speed of biochemical reactions but themselves remain unchanged.
• After completion of the reaction and release of the product they remain active to catalyse again.
• A small quantity of enzymes can catalyse the transformation of a very large quantity of the substrate into an end product.
• For example, sucrase can hydrolyse 1,00,000 times of sucrose as compared with its own weight.
• Specificity of action :
• The ability of an enzyme to catalyse one specific reaction and essentially no other is perhaps its most significant property.
• Each enzyme acts upon a specific substrate or a specific group of substrates.

• Reversibility of action :
• Enzymes are very sensitive to temperature and pH.
• Each enzyme exhibits its highest activity at a specific pH, called optimum pH.
• Any increase or decrease in pH causes decline in enzyme activity
• e.g. enzyme pepsin (secreted in stomach) shows highest activity at an optimum pH of 2 (acidic).
• Trypsin (in duodenum) is most active at an optimum pH of 9.5 (alkaline).
• Both these enzymes viz. pepsin and trypsin are protein digesting enzymes.

• Temperature :
• Enzymes are destroyed at higher temperature of 60-70°C or below, they are not destroyed but become inactive.
• This inactive state is temporary and the enzyme can become active at suitable temperature.
• Most of the enzymes work at an optimum temperature between 20°C and 35°C.

• Nomenclature of Enzymes :
• There are various ways of naming enzymes.
• Enzymes are named by adding the suffix-‘ase’ to the name of the substrate on which they act
• e.g.
• Protease- break up proteins
• Sucrase- break up sucrose
• Nuclease- break up nucleic acids
• The enzymes can be named according to the type of function they perform
• e.g.
• dehydrogenase remove hydrogen,
• carboxylase add CO;
• decarboxylases remove CO2,
• oxidases helping in oxidation.

• Some enzymes are named according to the source from which they are obtained
• e.g.
• Papain from Papaya,
• Bromelain from Pineapple, the member of Bromeliaceae family,.

• According to international code of enzyme nomenclature, the name of each enzyme ends with an -ase and consists of double name.
• The first name indicates the nature of substrate upon which the enzyme acts and the second name indicates the reaction catalyzed.
• e.g.
• Pyruvic decarboxylase catalyses the removal of CO₂ from the substrate pyruvic acid.
• Glutamate pyruvate transaminase catalyses the transfer of an amino group from the substrate glutamate to another substrate pyruvate.

• Pyruvic decarboxylase catalyses the removal of CO₂ from the substrate pyruvic acid.

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• Glutamate pyruvate transaminase catalyses the transfer of an amino group from the substrate glutamate to another substrate pyruvate.

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Classification of Enzymes :

• Oxidoreductases :
• These are enzymes catalyzing oxidation and reduction reactions by the transfer of hydrogen and/or oxygen.
• e.g. Alcohol dehydrogenase

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​

​

Alcohol

+

NAD ⁺

Alcohol

​

dehydrogenase

Aldehyde

+

NADH₂

• Transferases :
• These enzymes catalyse the transfer of certain groups between two molecules.
• e.g. Glucokinase

​

Glucose

+

ATP

Glucokinase

​

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Glucose-6-Phosphate

+

ADP

• Hydrolases:
• These are enzymes catalyse hydrolytic reactions.
• This class includes amylases, proteases, lipases etc.
• e.g. Sucrase

​

Sucrose

Sucrase

​

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Glucose

+

Fructose

• Lyases :
• These enzymes are involved in elimination reactions resulting in the removal of a group of atoms from substrate molecule to leave a double bond.
• It includes aldolases, decarboxylases, and dehydratases,
• e.g. Fumarate hydratase.

Histidine

Histidine

​

decarboxylase

Histamine

+

CO₂

• Isomerases :
• These enzymes catalyze structural rearrangements within a molecule.
• Their nomenclature is based on the type of isomerism.
• Thus these enzymes are identified as racemases, epimerases, isomerases, mutases,
• e.g. Xylose isomerase.

• Ligases or Synthetases :
• These are the enzymes which catalyse the covalent linkage of the molecules utilizing the energy obtained from hydrolysis of an energy-rich compound like ATP, GTP.
• e.g. Glutathione synthetase, Pyruvate carboxylase.

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Pyruvate + CO2 + ATP

Pyruvate carboxylase

Oxaloacetate + ADP + Pi

Mechanism of enzyme action :

• The basic mechanism by which enzymes catalyze chemical reactions begins with
• the binding of the substrate (or substrates) to the active site on the enzyme.
• The active site is the specific region of the enzyme which combines with the substrate.
• The binding of the substrate to the enzyme causes changes in the distribution of electrons in the chemical bonds of the substrate and ultimately causes the reactions that lead to the formation of products.
• The products are released from the enzyme surface to regenerate the enzyme for another reaction cycle.
• There are two models to explain the mechanism of forming Enzyme-Substrate complex, as described below:

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+

• Lock and Key model:
• The specific action of an enzyme with a single substrate can be explained using a Lock and Key analogy.
• This was first postulated in 1894 by Emil Fischer.
• In this analogy,
• the lock is the enzyme and
• the key is the substrate.
• Only the correctly sized key (substrate) fits into the key hole (active site) of the lock (enzyme).

• Induced Fit model (Flexible Model):
• Koshland (1959) proposed the induced fit theory.
• This states that approach of a substrate induces a conformational change in the enzyme.

• It is the more accepted model to understand mode of action of enzyme.
• Unlike the lock-and-key model,
• the induced fit model shows that enzymes are rather flexible structures in which the active site continually reshapes by its interactions with the substrate until the time the substrate is completely bound to it
• (it is also the point at which the final form and shape of the enzyme is determined).

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• Factors Affecting Enzyme Activity :
• Following factors affect enzyme activity :
• 1. Concentration of Substrate :
• Increase in the substrate concentration gradually increases the velocity of enzyme activity within the limited range of substrate levels.
• A rectangular hyperbola is obtained when velocity is plotted against the substrate concentration.
• Three distinct phases (A, B and C) of the reaction are observed in the graph.
• Where
• V = Measured velocity,
• V_max = Maximum velocity,
• S = Substrate concentration,
• K_m = Michaelis-Menten constant.

• K_m or the Michaelis-Menten constant is defined as the substrate concentration (expressed in moles/lit) to produce half of maximum velocity in an enzyme catalysed reaction.
• It indicates that half of the enzyme molecules (i.e. 50%) are bound with the substrate molecules when the substrate concentration equals the K_m value.

• K_m value is a constant and a characteristic feature of a given enzyme.
• It is a representative for measuring the strength of ES complex.
• A low K_m value indicates a strong affinity between enzyme and substrate, whereas a high K_m value reflects a weak affinity between them.
• For majority of enzymes, the K_m values are in the range of 10^-5 to 10^-2 moles.

• 2. Enzyme Concentration :
• The rate of an enzymatic reaction is directly proportional to the concentration of the substrate.
• The rate of reaction is also directly proportional to the square root of the concentration of enzymes.
• It means that the rate of reaction also increases with the increasing concentration of enzyme.
• And the rate of reaction can also decreased by decreasing the concentration of enzyme.

• 3. Temperature :
• The enzymatic reaction occurs best at or around 37^oC which is the average normal body temperature in homeotherms.
• The rate chemical reaction is increased by a rise in temperature but this is true only over a limited range of temperature.
• Enzymes rapidly denature at temperature above 40^oC.
• The activity of enzymes is reduced at low temperature.
• The temperature at which the enzymes show maximum activity is called Optimum temperature.

• 4. Effect of pH :
• Similar to temperature, there is also pH at which an enzyme will catalyze the reaction at the maximum rate.
• Every enzyme has different optimum pH value.
• The enzyme cannot perform its function beyond the range of its pH value.

• 5. Other Substances :
• The enzymes action is also increased or decreased in the presence of some other substances such as
• co-enzymes,
• activators and
• inhibitors.
• Most of the enzymes are combination of a co-enzyme and an apo-enzyme.
• Activators are the inorganic substances which increase the enzyme activity.
• Inhibitor is the substance which reduces the enzyme activity.

• Concept of Metabolism:
• Metabolism is the sum of the chemical reactions that take place within each cell of a living organism and provide energy for vital processes and for synthesizing new organic material.
• It involves continuous process of breakdown and synthesis of biomolecules through chemical reactions.
• Each of the metabolic reaction results in a transformation of biomolecules.
• Most of these metabolic reactions do not occur in isolation but are always linked with some other reactions.
• In living systems, cells are ‘work centres’ where metabolism involves two following types of pathways.

• a. Catabolic pathways:
• This lead to formation of simpler structure from a complex biomolecules.
• e.g. when we eat wheat, bread or chapati, our gastrointestinal tract digests (hydrolyses) the starch to glucose units with help of enzymes and releases energy in form of ATP (Adenosine triphosphate).
• b. Anabolic pathway:
• It is called biosynthetic pathway that involves formation of a more complex biomolecules from a simpler structure.
• e.g. synthesis of glycogen from glucose and protein from amino acids.
• These pathways consume energy.

• Metabolic pool :
• It is the reservoir of biomolecules in the cell on which enzymes can act to produce useful products as per the need of the cell.
• The concept of metabolic pool is significant in cell biology because it allows one type of molecule to change into another type
• e.g. carbohydrates can be converted to fats and vice-versa.
• Catabolic chemical reaction of glycolysis and Krebs cycle not only provide ATP but also makes available metabolic pool of biomolecules that can be utilized for synthesis of many important cellular components.
• The metabolites can be added or withdrawn from this pool according to the need of the cell.
• The balance between catabolism and anabolism maintain homeostasis in the cell as well as in the whole body.

• Secondary metabolites (SMs) :
• Secondary metabolisms are small organic molecules produced by organisms that are not essential for their growth, development and reproductions.
• Several types of bacteria, fungi and plants produce secondary metabolites.
• Secondary metabolites can be classified on the basis of
• Chemical structure
• e.g. SMs containing rings, sugar,
• Composition (with or without nitrogen),
• Their solubility in various solvents, or
• The pathway by which they are synthesized
• e.g. phenylpropanoid produces tannins.

• A simple way of classifying secondary metabolites includes three main groups such as,
• 1. Terpenes :
• Made from mevalonic acid that is composed mainly of carbon and hydrogen.
• 2. Phenolics :
• Made from simple sugars containing benzene rings, hydrogen and oxygen.
• 3. Nitrogen- congaing compounds :
• Extremely diverse class may also contain sulphur.

Economic importance of Secondary Metabolites -

1. Secondary metabolites from natural sources have made a significant contribution for millennia.
• In modern medicine, drugs developed from secondary metabolites have been used to treat infectious diseases, cancer, hypertension and inflammation.
2. Morphine was the first alkaloid isolated from plant Papaver somniferum. (अफू)
• It is used as pain reliever and cough suppressant.
3. SMs like alkaloids nicotine and cocaine and the terpenes, cannabinol are widely used for recreation and stimulation.
4. Flavours of secondary metabolites improve our food preference.
5. Characteristic flavours and aroma of cabbage and its relatives are caused by nitrogen and sulphur-containing chemicals, glucosinolates, protect these plants from many pests.
6. Tannins are added to wines and chocolate for improving astringency. (तुरटपणा)
7. Since most of secondary metabolites are having antibiotic properties, they are also used as food preservatives.