Enzymes ~

Supermolecule
 Why Super molecule ?
The superior designs of biological super molecules
become apparent only upon studying them
Information Conversion and Amplification:
Signal Transduction
When an extracellular signal is recognized by a
receptor on a cell membrane, the G-protein
activates the enzyme inside the cell. The
activation of an enzyme by external chemicals can
be mimicked using a system consisting of an
artificial receptor and an enzyme immobilized on
an artificial lipid bilayer membrane.
2
 Energy Conversion –
Photosynthesis
▸ During the process of photosynthesis, a
well-organized dye array in the cell
membrane accomplishes photoinduced
charge separation that eventually leads
to ATP synthesis. This system has been
mimicked by immobilizing a functional
dye molecule and ATP synthase in a
lipid bilayer membrane.

3
Material Conversion –
Natural and Artificial Enzymes
▸ Enzymes perform highly selective
and highly efficient molecular
conversion based on sophisticated
three-dimensional arrangements of
amino acids. Artificial enzyme
mimics can be constructed using First artificial enzyme created with two
cyclodextrins and lipid bilayer non-biological groups
membranes
4
 Cleaving Genes –
Restriction Enzymes
▸ Restriction enzymes can cleave
nucleic acids at specific
sequences. Artificial restriction
enzymes can be prepared by
combining oligo (nucleic acids)
and supramolecular catalytic
sites.
5
 Tailor-Made Enzymes –
Catalytic Antibodies
▸ There are antibodies that
catalyze reactions by
mimicking their transition
states. A catalytic antibody
can be regarded as a tailor-
made artificial enzyme.
6
 Key to the Origin of Life –
Ribozymes
▸ Ribozymes are nucleic acids
with catalytic capabilities. The
discovery of ribozymes led to
the RNA world hypothesis for
the origin of life.

7
Quiz
How many enzymes are there in human body?
 75000 Enzymes in human body and 1300 enzymes in

“
single cell.
▸ Examples of digestive enzymes are:
▸ Amylase, produced in the mouth. It helps break down large starch
molecules into smaller sugar molecules.
▸ Pepsin, produced in the stomach.
▸ Trypsin, produced in the pancreas.
▸ Pancreatic lipase, produced in the pancreas.
▸ Deoxyribonuclease and ribonuclease, produced in the pancreas.
9
 Enzyme FAQ

▸  Vegetarian Enzymes, "The ideal" enzyme

source?

10
 Timeline

11
 Industry Standards for evaluation of plant & vegetarian enzymes
▸When comparing enzyme products make sure enzyme
activities are measured using units such as these:
Alpha-Galactosidase - GALU (Galactosidase
Units)
Amylase - DU (Dextrinizing Units)
Bromelain - GDU (Gelatin Digesting Units) or
FCCPU
Catalase - Baker Units (Named after the Author)
Cellulase - CU (Cellulase Unit)
Glucoamylase - AGU (Amyloglucosidase Units)
Hemicellulase - HCU (Hemicellulase Units)
Invertase - INVU (Invertase Activity Unit) or
SU (Sumner Units)
Lactase - ALU (Acid Lactase Unit)
12
Lipase - FCCFIP (Federation
Internationale Pharmceutique)
Maltase - DP (Degrees of Diastatic
Power)
Nattokinase - FU (Fibrinolytic
Units)
Pectinase - Endo-PGU (Endo-
Polygalacturonase units)
Phytase - FTU (Phytase Units)
Protease-HUT (Hemoglobin Unit
on a L-Tyrosine basis)
Xylanase - XU (Xylanase Units)
 Activity: 1 E
▸ Proceed for extraction of Enzyme
Papain
▸ Write a step wise protocol
▸ Prepare a trip plan to visit papain
extraction plant.
▸ Submission date: August 14

13
 Enzymes : Sources
•

14
 Sources

15
 Sources

16
 EXAMPLES
substrate enzymes products
lactose lactase glucose + galactose
maltose maltase Glucose
cellulose cellulase Glucose
lipid lipase Glycerol + fatty acid
starch amylase Maltose
protein protease Peptides +
polypeptide
 CLASSIFICATION OF ENZYMES
 A systematic classification of enzymes has been developed by
International Enzyme Commission.

 This classification is based on the type of reactions catalyzed

by enzymes.

 There are six major classes.

 Each class is further divided into sub classes, sub sub-classes

and so on, to describe the huge number of different enzyme-
catalyzed reactions.
 Classification of enzymes Continued……..

ENZYME CLASS REACTION TYPE EXAMPLES

Oxidoreductases Reduction-oxidation Lactate
(redox) dehydrogenase
Transferases Move chemical group Hexokinase
Hydrolases Hydrolysis; bond cleavage Lysozyme
with transfer of functional
group of water

Lysases Non-hydrolytic bond Fumarase

cleavage
Isomerases Intramolecular group Triose phosphate
transfer (isomerization) isomerase

Ligases Synthesis of new covalent RNA polymerase

bond between substrates,
using ATP hydrolysis
Examples in Metabolism
Examples are dehydrogenases catalyse reactions in which a substrate is
oxidised or reduced
Transaminases which catalyze the transfer of amino group or kinases which
catalyze the transfer of phosphate groups.

Lipases catalyze the hydrolysis of lipids, and proteases catalyze the hydrolysis of
proteins
Decarboxylases catalyze the removal of carboxyl groups
Isomerases may catalyze the conversion of an aldose to a ketose, and mutases
transfer functional group from one atom to another within a substrate.

Synthetases link two smaller molecules are form a larger one.

 Mechanism of Enzyme action
Energetics of enzymatic reactions
Conversion of substrate to
product proceeds through
formation of a
transition state.
The free energy of activation of an uncatalyzed
reaction is very high. Enzymes form favorable
interactions with the substrate and facilitate
formation of the transition state by lowering the
free energy of activation. The transition state then
dissociates to give the product and regenerates free
enzyme. For a reaction to be spontaneous, the ∆G
must be negative. It must be emphasized that
enzymes do not alter the equilibrium of a reaction.
 Enzymes as a Desired Product
Enzyme Industry Application

Isopropyl myristate production
Novozym-435 Consumer Goods
(Cosmetic)

Bromelain Food Meat tenderizer

Noopazyme Food Improve noodle quality

Asparaginase Pharmaceutical Lymphatic cancer therapeutic

Ficin Pharmaceutical Digestive aid

Urokinase Pharmaceutical Anticoagulant

β-Lactamase Pharmaceutical Penicillin allergy treatment

Subtilisin Consumer Goods Laundry detergent

Name any industry that produces enzymes

AB Enzymes develops enzyme products for food, animal feed

and technical applications.
 Enzyme in industry
▸ The great majority of microbial enzymes come from a very limited number of
genera, of which Aspergillus species, Bacillus species and Kluyveromyces (also
called Saccharomyces) species predominate.
▸ Most of the strains used have either been employed by the food industry for
many years or have been derived from such strains by mutation and selection.
▸ There are very few examples of the industrial use of enzymes having been
developed for one task.
▸ Shining examples of such developments are the production of high fructose
syrup using glucose isomerase and the use of pullulanase in starch hydrolysis.

24
 Commercialization
▸ Producers of industrial enzymes and their customers will share the common aims of
economy, effectiveness and safety.
▸ They will wish to have high-yielding strains of microbes which make the enzyme
constitutively and secrete it into their growth medium (extracellular enzymes).
▸ If the enzyme is not produced constitutively, induction must be rapid and
inexpensive.
▸ Producers will aim to use strains of microbe that are known to be generally safe.
Both producers and users will wish to have the enzymes in forms that present
minimal hazard to those handling them or consuming their product.

25
 Specialised business
▸ The development of commercial enzymes is a specialised business which is
usually undertaken by a handful of companies which have high skills in
▸ screening for new and improved enzymes,
▸ fermentation for enzyme production,
▸ large scale enzyme purifications,
▸ formulation of enzymes for sale,
▸ customer liaison, and
▸ dealing with the regulatory authorities.

26
 Apparatus for enzyme extraction

27
28
 Characteristics of Enzyme
 Enzymes speed up the reaction by lowering the activation energy
of the reaction.
 Their presence does not effect the nature and properties of
end product.
 They are highly specific in their action that is each enzyme
can catalyze one kind of substrate.
 Small amount of enzymes can accelerate chemical reactions.
 Enzymes are sensitive to change in pH, temperature and substrate
concentration.
 Turnover number is defined as the number of substrate
molecules transformed per minute by one enzyme molecule.
Catalase turnover number = 6 x106/min
29
 Enzyme assay

The biochemist needs a test, called an assay, for some unique identifying property of the protein so
that he or she can tell when the protein is present. Determining an effective assay is often difficult; but
the more specific the assay, the more effective the purification.

Ability to absorb light at particular nm

Consequently, we can follow the progress of the reaction by examining how much light the reaction
mixture absorbs at 340 nm in unit time—for instance, within 1 minute after the addition of the
enzyme. Our assay for enzyme activity during the purification of lactate dehydrogenase is thus the
increase in absorbance of light at 340 nm observed in 1 minute.
To be certain that our purification scheme is working, we need one additional piece of information—
the amount of protein present in the mixture being assayed.
There are various rapid and accurate means of determining protein concentration. With these
two experimentally determined numbers—enzyme activity and protein concentration—we then
calculate the specific activity, the ratio of enzyme activity to the amount of protein in the
enzyme assay. The specific activity will rise as the purification proceeds and the protein mixture
being assayed consists to a greater and greater extent of lactate dehydrogenase. In essence, the
point of the purification is to maximize the specific activity.

30
 Thanks!
Any questions?
You can find me at:
pragyasrathore@gmail.com
9425070708

31
 References
https://www.eurekalert.org/pub_releases/2020-02/uog-fae020720.php

https://www.sciencedirect.com/topics/biochemistry-genetics-and-molecular-biology/artificial-enzyme

https://www2.chemistry.msu.edu/courses/cem958/FS04_SS05%5Cfrawley.pdf

https://www.rndsystems.com/research-area/proteases---other-enzymes

https://www.sciencedirect.com/science/article/abs/pii/S0920586118317218

https://www.eurekaselect.com/169519/article

32
Thank you