Enzymes – Definition, properties, classification, applications & Isolation

Definition, properties, classification, applications & Isolation

Enzymes Definition, properties, classification, applications & Isolation

Session Objectives

At the end of the session, student will be able to

Ø    Describe the properties of enzymes

Ø    Classify enzymes

Ø    Discuss the various applications of enzymes

Ø    Explain the different steps involved in isolation of enzymes


Ø  Enzymes – soluble, colloidal organic catalysts – produced by living cells but are capable of acting independently of the cells

Ø  Accelerates and catalyses thousands of biochemical reactions in the living cells   

Ø  Functional unit of cell metabolism 

Ø  Proteinaceous in nature

Properties of Enzymes

     Proteinaceous and High molecular weight – 12,000 – 1 million

     Colloidal in nature

      Soluble – water, buffer and dilute alcohol

       Insoluble – Conc. Alcohol, acetone and other organic solvent

      Highly selective and specific  – action

      Required in small quantities relative to the substrate concentration

      35˚ C  – 40˚ C

      Temp above 65˚ C or 0˚ C – loss of catalytic activity – due to the disruption of polypeptide chains

      Hence stored – low temp  – moisture free container

      Enzymes – require – a specific, heat stable, low molecular weight organic molecule – Coenzyme

      Some enzymes – coenzyme and one more metal ions  – activity

      Does not change the final equilibrium position of reactions – only the rate of attainment of equilibrium – increased

Non-protein part – coenzyme or metal (Prosthetic group) + Protein part (Apoenzyme) = Enzymes

Structure of enzyme

       Co enzymes – Coenzyme A, FAD, NAD

       Co factors – Glucose – 6-phosphate – Mg, Arginase – Mn

       Protein – Linear chain of amino acid residues joined  – peptide bond

       Catalytic activity depends  – L-amino acid sequence and peptide bond constituting the protein molecule

       Localized folding of primary structure – secondary structure

       Overall folding – molecule – tertiary structure

       Agglomeration of several folded chains – Quaternary structure

Classification of Enzymes

IUB classification:

1. Oxido reductases – catalyzing oxidation and reduction between two substrates 

S reduced + S’ oxidized = S oxidized + S’ reduced

Eg. Alcohol dehydrogenase, lactate dehydrogenase

2. Transferase – catalyzing – transfer of S, C, N, P other than H

S – G + S’ = S’- G +S

Eg. Hexokinase, acetyl transferase

3. Hydrolases – catalyzing – hydrolysis of ester, ether, peptides, glycosyl, C-C, C-halide bonds

Acylcholine + H2O = Choline + acid

Eg. Urease

4. Lyases – Catalyses removal of groups from substrates – other than hydrolysis leaving double bond – acts on C-C, C-O, C-N, C-S bond

Eg.  Aldolase, fumerase

5. Isomerase – catalyses – interconversion – optical, geometric or positional isomers —- Eg. Alanine isomerase

6. Ligases – an enzyme that can catalyze the joining of two large molecules by forming a new chemical bond, usually with accompanying hydrolysis of chemical group on one of the larger molecules. Eg. Pyruvate carboxylase, glutamine synthetase                                           

Other methods of classification

1. Extracellular enzymes or exoenzymes  – secreted outside the cell

Eg. Cellulose, polyglucturonase

2. Intracellular enzymes or endoenzymes – secreted within the cell

Eg. Invertase, asparaginase

Applications of Enzyme:

Medicinal applications

Digestive disorders-

         Papain (Papaya)

         Pancreatin (animal pancreas)

         ß-Galactosidase (A. Oryzae)

         Penicillinase (B. subtilis)

Deworming agents

         Papain (Papaya)

         Ficin (fig)

Anticancer agents

         Asparginase (E coli, guinea pig)


         Bromelain (Pineapple)

Anti-coagulant agents

         Streptokinase (β-hemolytic streptococci)

         Urokinase (Human urine)

Surface disinfectants

         Trypsin (animal pancreas)

Diagnostic agents

         Glucose isomerase (diabetes)

         SGOT, SGPT, ALP (Liver disorders)

Upper Respiratory disorders

         Chymotrypsin (Bovine pancreas)

Food industrial applications

Tenderization of meat

         Papain (Papaya)

         Bromelain (Pineapple)

Ice cream industry

         Lactase (prevention of lactose crystals)

Beverage industry

         Invertase (yeast)

Chocolate industry

         Invertase (yeast)

Juice & wine processing


Industrial applications

Textile industry (Destarching)

         Amylase (Bacteria, fungi)

Leather industry (Bating)

         Proteolytic enzymes (Bacteria, and fungi)

Detergents (Destaining)


Paper industry (Bleaching of pulp)

         Xylenase (Bacteria)

         Cellulase (Bacteria)

Organic compounds


Enzymes – Production

Source of Enzymes: Living cells

1.  Plant:
Papain, Bromelain

2.  Animal: Urokinase

3.  Microorganism:

Advantages of microorganisms over other sources of enzymes

Ø    Growth is very fast – Grown on medium containing cheap raw materials

Ø   Genetic engineering and manipulations of microbial cells – possible – increase the yield of enzyme

Ø   Large quantities of enzymes are produced

Ø   Animal sacrifice can be minimized

Selection of microorganisms:

Ø     Non-pathogenic

Ø    Should produce extracellular enzymes

Ø    Fermentation time should be less

Ø    Must grow on medium with cheap raw material

Ø    Must give a high yield of enzymes

Ø    Must be compatible with physical and chemical properties of the medium

Ø    Strain must not produce byproducts – inhibit the growth of m/o  Aspergillus and Bacillus – industrial production

Isolation of enzymes

1. Extraction                            

Cell disintegration/ Cell disruption

Removal of lipids

2. Preparation of crude enzyme

Removal of nucleic acids


Addition of acid or base

3. Precipitation

Salting out method

Addition of organic solvents

Addition of non-ionic polymers


4. Purification

Chromatographic techniques


1. Extraction

Ø    Liberation of enzymes from the cells or cellular constituents

Ø    Break the cell wall or membrane – physical, chemical, mechanical

Ø    Extraction medium: Buffer solution, Temperature, and pH : Optimum

Modification of extraction medium: To achieve enzyme with maximum activity buffers can be modified by adding

              EDTA (To remove heavy metals)

              Mercapto ethanol (to prevent the breakage of disulfide bond of enzymes containing cysteine amino acid)

              Triton-X (cell disruption)

Disintegration of animal and plant tissues

             Plant tissues: By using a hammer mill/chopper mill

             Animal tissues:   Organs or muscles        



Minced – vertical cutter mixer


Colloidal mill


Cell disintegration


Ø  Enzymes are biocatalysts, high mol.wt, Proteinaceous and water-soluble compounds

Ø    Activity  is affected by temp, pH, and heavy metals, specific in their action

Ø     Classification according to IUMAB and site of action

Ø     Enzymes have medicinal, food, and industrial applications

Ø     Main source is plant, animal, and microorganisms

Ø     Isolation involves extraction, preparation of crude enzyme, precipitation, and purification

Also, Visit: Biotechnology Notes

B Pharma Notes

Enzyme Notes