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Enzymes

Enzymes are proteins that catalyze (i.e. speed up) biochemical reactions and are not changed during the reaction. 

Substrate

In enzymatic reactions, the molecules at the beginning of the process are called substrates, and the enzyme converts them into different molecules, the products. Almost all processes in a cell need enzymes in order to occur at significant rates.

Characteristics of enzymes

There are four characteristics of enzyme action
1- Factors affecting 
2-The rate of enzyme action
 3-Mechanism of enzyme action 
 4-Specificity of enzymes

Metablism

 The term metabolism is derived from a Greek word meaning "change". The concept of metabolism was first of all given by Ibn-e-Nafees, who stated that "the body and its parts are always undergoing change". The life of living organisms is a reflection of what is going on in their bodies. 
Metabolism is the set of biochemical reactions that occur in living organisms in order to maintain life. These processes allow organisms to grow and reproduce, maintain their structures, and respond to their environments. Biochemical reactions in living organisms are essentially energy transfers.

Anabolism

 Anabolism is the total series of chemical reactions involved in the synthesis of compounds. 

Catabolism

Catabolism is the series of chemical reaction. During catabolism chemicals are transformed from one form to the other by enzymes. Enzymes are crucial to catabolism because they act as biocatalysts and speed up and regulate the metabolic pathways .

Activation energy and enzymes

 All chemical reactions require activation energy to break chemical bonds and begin the reaction. The need for activation energy acts as a barrier to the beginning of reaction . Enzymes lower such barriers by decreasing the requirement of activation energy. Thus, in the presence of enzymes, reactions proceed at a faster rate. Enzymes lower the activation energy in several ways. 
They do so by Altering the shape of the substrates and reducing the amount of energy required to complete the transition Disrupting the charge distribution Bringing substrates in the correct orientation to reaction Progress of reaction All biochemical catalysts are not proteins, for example some RNA molecules also catalyze reactions. In 1878, German physiologist Winhelm Kuhne first used the term enzyme. 
Enzymes are globular proteins and range from 62 to over 2,500 amino acids. Like all proteins, enzymes are made of long, Most enzyme reaction rates are millions of times faster than those of comparable uncatalyzed reactions. As with all catalysts, enzymes are not consumed by the reactions they catalyze. 
 Enzymes are usualy very specific for the type of reaction and for the nature of the' substrates. linear chains of amino acids that fold to produce a three-dimensional molecule 
Almost all enzymes are proteins i.e. they are made of amino acids. The activities of enzymes are determined by a small portion of the enzyme molecule (around 34 amino acids) which is directly involved in catalysis. This catalytic region, known as the active site, recognizes and binds the substrate, and then carries out the reaction. 
Since enzymes are extremely selective for their substrates and speed up only a few reactions, the set of enzymes made in a cell determines which metabolic pathways occur in that cell. 

Active sites of enzymes

Enzymes can be categorized on the basis of the site where they work i.e. they may be intracellular enzymes (e.g. enzymes of glycolysis working in the cytoplasm) or may be extracellular enzymes (e.g. pepsin enzyme working in the stomach cavity). 
 The enzyme activity is controlled in the cell by many ways. Enzyme production can be enhanced or diminished by a cell in response to changes in the cell's environment. Enzyme activity can also be regulated by inhibitors and activators. Some enzymes do not need any additional components to show full activity.
 However, others require non-protein molecules or ions called cofactors for activity. Cofactors can be either inorganic (e.g. metal ions) or organic (e.g. flavin and heme). If organic cofactors are tightly bound to enzyme they are called prosthetic groups, but if they are loosely attached with enzyme, they are called coenzymes. Coenzymes are small organic molecules that transport chemical groups from one enzyme to another. 
Some important coenzymes are vitamins (e.g. riboflavin, thiamine and folic acid). 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. 
Enzymes are extensively used in different industries for fast chemical reactions. For example; Food industry: Enzymes that release sugar molecules from starch are used in production of white bread, buns, and rolls. Enzymes are also used for the production of cheese. , Brewing industry: Enzymes degrade starch and proteins to produce simple sugars and amino acids that are used by yeast for fermentation (to produce alcohol).

Role of enzymes in paper industry

Enzymes degrade starch to lower its viscosity that aids in making paper. Biological detergent: Protease enzymes are used for the removal of protein stains from clothes. Amylase enzymes are used in dish washing to remove resistant starch residues.
 complete the transition Disrupting the charge distribution Bringing substrates in the correct orientation to react All biochemical catalysts are not proteins, for example some RNA molecules also catalyze reactions. 
In 1878, German physiologist Winhelm Kuhne first used the term enzyme. Enzymes are globular proteins and range from 62 to over 2,500 amino acids. Like all proteins, enzymes are made of long, Most enzyme reaction rates are millions of times faster than those of comparable uncatalyzed reactions. 
As with all catalysts, enzymes are not consumed by the reactions they catalyze. Enzymes are usualy very specific for the type of reaction and for the nature of the' substrates. linear chains of amino acids thatfold to produce a three-dimensional molecule 
Active site
 Almost all enzymes are proteins i.e. they are made of amino acids. The activities of enzymes are determined by a small portion of the enzyme molecule (around 34 amino acids) which is directly involved in catalysis. This catalytic region, known as the active site, recognizes and binds the substrate, and then carries out the reaction. 

 Since enzymes are extremely selective for their substrates and speed up only a few reactions, the set of enzymes made in a cell determines which metabolic pathways occur in that cell. Enzymes can be categorized on the basis of the site where they work i.e. they may be intracellular enzymes (e.g. enzymes of glycolysis working in the cytoplasm) or may be extracellular enzymes (e.g. pepsin enzyme working in the stomach cavity). 
 The enzyme activity is controlled in the cell by many ways. Enzyme production can be enhanced or diminished by a cell in response to changes in the cell's environment. Enzyme activity can also be regulated by inhibitors and activators. 

Co-factors

 Some enzymes do not need any additional components to show full activity. However, others require non-protein molecules or ions called cofactors for activity. 
Cofactors can be either inorganic (e.g. metal ions) or organic (e.g. flavin and heme). If organic cofactors are tightly bound to enzyme they are called prosthetic groups, but if they are loosely attached with enzyme, they are called coenzymes. Coenzymes are small organic molecules that transport chemical groups from one enzyme to another. 

Important co-enzymes

Some important coenzymes are vitamins (e.g. riboflavin, thiamine and folic acid). 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.
 Enzymes are extensively used in different industries for fast chemical reactions. 

Use of enzymes in food industry 

Enzymes that release sugar molecules from starch are used in production of white bread, buns, and rolls. Enzymes are also used for the production of cheese.

Use of enzymes in Brewing industry: 

Enzymes degrade starch and proteins to produce simple sugars and amino acids that are used by yeast for fermentation (to produce alcohol). 

  Use of enzymes in Paper industry: 

Enzymes degrade starch to lower its viscosity that aids in making paper

Enzymes as Biological detergent: 

Protease enzymes are used for the removal of protein stains from clothes. Amylase enzymes are used in dish washing to remove resistant starch residues. 

Factors affecting the rate of enzyme action

 Enzymes are very sensitive to the environment in which they work. 
The activity of an enzyme is affected by any change in conditions that alters its chemistry and its shape.
 Some of the factors that can affect the rate of enzyme action are being discussed below. 

 Effect of temperature on enzymes

Increases in temperature will speed up the rate of enzyme catalyzed reactions, but only to a point .
 Every enzyme works at its maximum rate at a specific temp erature for § temperature called as the optimum temperature for the maximum working speed that enzyme. 
When temperature rises to a certain limit, of  human enzymes is 37°C. the heat adds in the activation energy and also provides kinetic energy and so reactions are accelerated. But when temperature is raised well above the optimum temperature, the heat energy increases the vibrations of atoms of enzyme molecules and the globular structure of enzyme is lost. 
This is known as denaturation of enzyme. It results in a rapid decrease in the rate of enzyme action and it may be blocked completely Substrate concentration: If there are enzyme molecules with vacant active sites, an increase in substrate concentration will increase the rate of reaction.

Saturation of active sites of substrate

 If the enzyme concentration is kept constant and the amount of substrate is increased, a point is reached where any further increase in substrate does not increase the rate of reaction any more. When all the active sites of the enzymes are occupied (at high substrate concentration), any more substrate molecules do not find free active sites. This state is called saturation of active sites and reaction rate does not increase .

 Optimum pH

 All enzymes work at their maximum rate at a narrow range of pH, called as the optimum pH . A slight change (increase or decrease) in this pH causes retardation in enzyme activity or blocks it completely. Every enzyme has its specific optimum pH value. For example pepsin (working in stomach) is active in acidic medium (low pH) while trypsin (working in small intestine) shows its activity in alkaline medium (high pH). 
Change in pH can affect the ionization of the amino acids at the active sites.

 Enzyme -Substrate complex


 When enzyme attaches with the substrate, a temporary enzyme-substrate (ES) complex is formed. The enzyme catalyzes the reaction and substrate is transformed into product. The ES complex breaks and enzyme and product are released.

 Enzyme action

 Enzyme action In order to explain the mechanism of enzyme action a German chemist Emil Fischer, in 1894, proposed the lock and key model. According to this model both the enzyme and the substrate possess specific complementary geometric shapes that fit exactly into one another. This model explains enzyme specificity.

 Lock and key model

 The “induced fit model” is more acceptable than the “lock and key” model.
 In 1958 an American biologist Daniel Koshland suggested a modification to the lock and key model and proposed the induced-fit model. He said that enzymes are flexible structures and their active site is reshaped as the substrate interacts with the enzyme. According to this model the active site is not a rigid structure rather it is molded into the precise position to Perform its function. Substrate. 

Specificity of enzymes

 There are over 2000 known enzymes, each of which is involved in one specific chemical reaction. Enzymes are also substrate specific. The enzyme protease (which breaks peptide bonds in proteins) will not work on starch (which is broken down by an enzyme amylase). Similarly lipase enzyme acts only on lipids and digests them into fatty acids and glycerol.
 The specificity of different enzymes is determined by the shapes of their active sites. The active sites possess specific geometric shapes that fit with specific substrates.
Specificity of enzymes is  due to the geometric shape of active site Experiment to observe the invitro enzyme activity
We can design an experiment to observe the in-vitro enzyme activity. For this purpose we would select meat proteins as substrate and pepsin as the protein digesting enzyme. 

Apparatus

Meat, Test tube, pepsin solution, HCI, Biuret reagent 

Background information: 

 Animal flesh (meat) contains lot of proteins. Pepsin enzyme is produced in stomach (in its inactive form pepsinogen). It acts on protein molecules and digests them to peptides procedure: 

Procedure

Take a small piece of meat in two test tubes and pour 15 ml of pepsin in one of them the and pour 15 ml water in the second tube (for comparison).
 2. Add 10 drops of HCI in both test tubes and place them at 37°C in incubator. 

Observations

Observe the piece of meat after four hours. Perform the Biuret test to confirm the presence of proteins in both tubes. Results: The Biuret test gives negative results in the tube in which pepsin was added. It confirms that no proteins are present in this test tube and all have been digested by the enzyme pepsin. 

Evaluation: 

 Breakdown of the polysaccharide starch to the disaccharide maltose. It is present in saliva, plant tissues, and also in seeds. To observe the in-vitro enzyme activity we can also select starch as substrate and amylase as the starch digesting enzyme

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