Enzymes
What are Enzymes?
Enzymes are proteins that function as biological
catalysts. A catalyst is a substance that increases the
rate of a chemical reaction and is not changed by the
reaction. These enzymes are made in living cells and
thus are known as biological. From this information, we
can state that:
Enzymes are made in the body to speed up reactions
that take place in the body without getting changed or
used up.
As enzymes are made up of proteins, they will need
amino acids as building blocks for them.
The substrate (molecule on which the enzyme acts
upon) is the key to a lock (the enzyme). The whole
idea of the lock and key mechanism is known as the
lock and key hypothesis derived by the German
scientist Emil Fischer.
In the presence of the enzyme, the substrate attaches
to the enzyme and will either break (catabolic) or build
up (anabolic) into products. There may be multiple
substrates or products or even 1 of each. The product
will separate from the enzyme. If 2 products are formed
(such as fatty acids and glycerol broken down from fats
by lipase) the products will also separate from each
other.
Almost all metabolic processes (catabolic – breaking
down, and anabolic – building up processes) require
enzymes.
Enzymes and Metabolism
Metabolism is the sum of all catabolic and anabolic
processes that take place in a living body.
Catabolic
Enzymes break complex
substrates to simple
products
These reactions break
down and release energy
(exothermic)
Respiration is an
example: breaks down a
complex sugar (glucose)
into simpler compounds
and releases energy
Anabolic
Enzymes synthesize
complex products from
simple substrates
These reactions build up
and use energy
(endothermic)
Photosynthesis is an
example: takes in energy
from sunlight and
produces a complex
sugar (glucose)
The production of enzymes will be anabolic as they are
built from amino acids in cells.
Where are Enzymes Produced?
All enzymes are produced within cells. Most are
intracellular: work within the cell but some are
extracellular: released from cells and work outside
these cells (example: digestive enzymes such as
lipase).
Enzyme Action
Each enzyme is specific to a substrate (protease is
specific to proteins).
An enzyme is a 3D protein molecule with an active site
which allows the enzyme to act as a catalyst. The active
site is the point at which a substrate attaches to an
enzyme.
Enzymes are specific to one particular substrate as the
enzyme is a complementary shape to the substrate.
When the substrate moves into the enzyme’s active
site, they become known as the enzyme-substrate
complex. After the reaction has occurred, the products
leave the enzyme’s active site as they no longer fit it
and it is free to take up another substrate.
Enzymes are specific due to the shape of its active site.
After the reaction takes place, the enzymes remain
unchanged as they are catalysts.
The diagram below shows the action of both anabolic
and catabolic enzyme action:
Note that anabolic reactions are also known as
synthesis (condensation) reactions and catabolic
reactions may be known as hydrolysis if water is
involved in the breakdown.
Factors Affecting Enzyme Activity
Temperature:
 When the temperature is lower than the
optimum (ideal) temperature, fewer collisions
of enzymes and substrates occur as there is
lesser energy in the molecules.
 As the temperature increases, substrate
molecules move faster and collide with greater
energy, making it more likely for binding at the
active site.
 The optimum temperature is where enzymes
work best as most collisions are successful.
 At high temperatures, there is more energy, but
the active site starts to deform due to
vibrations, so fewer collisions take place (losing
ability to bind). This is called the denaturing of
enzymes.
The optimum temperature for enzymes to work in:
 Humans: 35 to 40°C – below this, works slow;
above this, denaturing takes place.
 Plants: 25 to 30°C – below this, works slow;
above this, denaturing takes place.
This can be plotted on a graph:
Power of Hydrogen (pH):
Summary of Enzyme Action:
1) Enzymes and substates randomly move about in
solution
2) When an enzyme and its complementary substrate
randomly collide – with the substrate fitting into the
active site of the enzyme – an enzyme-substrate
complex forms, and the reaction occurs.
3) A product forms from the substrate which is then
released from the active site. The enzyme is
unchanged and will go on to catalyse further
reactions.
The above scale is known as the pH Scale. It shows
the levels of acidity or how alkaline a substance is.
Most enzymes work best around neutral (6.5 to 8) pH,
though there are some which need acidic or slightly
alkaline conditions. Hydrochloric acid is produced in
the stomach for the enzyme pepsin (a protease) to
work at optimum ph. This acid will pass through the
intestine, thus hydrogen carbonates (alkaline) are
produced in the pancreas and passed on to make sure
the pH in the intestine is neutralized and to give
amylase and lipases its ideal pH.
When it comes to pH, enzymes only work at its
optimum pH and will be denatured below or above
optimum as the active sites shape gets changed and
then fewer collisions take place.
Biology in Industries
Ammonifying bacteria can process ammonia at
regular temperature and pressure (RTP) rather than
using Haber’s Process of producing ammonia which
requires a higher RTP.
Enzymes are biocatalysts and will work at regular RTP
and pH as found in living organisms. Therefore:
 If enzymes are used for industrial processes such as
making ammonia, then extreme and expensive
conditions won’t be needed.
 Enzymes can be obtained by microbes.
Note that the pH of the enzyme is not 2 (pepsin) or 7.5
(amylase), but it is the best pH it works at.
Note: Microbes secrete enzymes on substrates during
extracellular digestion. Using this knowledge,
scientists have created a device known as a biofermenter or bioreactor to obtain enzymes and maybe
ammonia from microbes as they secrete them.
The diagram below shows a bioreactor:
The parts of a bio-fermenter have specific uses:
Activators and Inhibitors:
These are molecules that affect how an enzyme binds
to a substrate:


Activators or coenzymes make binding of
enzymes more likely (Chlorine ions are present
and needed along with salivary amylase).
Inhibitors make it harder for enzymes to bind
to each substrate (Cyanide ions are considered
as poisonous as they block the active sites of
enzymes involved in respiration which inhibits
this process and can lead to death).
 Microbe inlet/input: The organisms that will carry
out fermentation (anaerobic) or other aerobic
processes are inputted from here.
 Nutrient inlet/input: The microbes require energy
(carbohydrates) and growth materials (amino acids)
which are inputted here.
 Paddle stirrer: They continuously mix the contents
of the bioreactor so that: microbes and nutrients are
always in contact; ensures even temperature
throughout and for aerobic reactions the mixing
occurs from an airstream which also provides
oxygen.
 Hot or Cool Water jacket: This maintains the
temperature all around the reactor so that enzymes
work at its best conditions. The temperature remains
constant all throughout.
 Gas outlet: Carbon dioxide and other gases may be
produced from respiration. These may be harmful for
the bacteria so there is a gas outlet which allows the
gas to escape and to avoid pressure building up in
the reactor. If this gas is carbon dioxide, then it may
be collected and used to make fizzy drinks.
 Probes (Sensors): They monitor conditions in the
reactor as they are connected to computers/
monitors. They will adjust the conditions if there are
any non-suitable conditions/ non-optimum
conditions.
 Further processing outlet: Further processing may
be necessary to separate microbes from the food/
enzymes and the microbes may be reused or
returned for the process to occur once again; and
preparing products for sale (involves drying or
crystallization). Sometimes, a tap may be present to
control the flow of output.
In a bioreactor, the culture (nutrients and microbes that
are inputted) must be pure (1 microbe type only) and
the equipment and nutrients must be sterile (cleaned)
to avoid competition for expensive nutrients (if any other
microbe manages to get inside) and to reduce diseases
(or spreading of toxic waste products of microbes which
causes microbes to contaminate the product –
decrease in yield).
Continuous
Culture:
In this,
microbes and
nutrients are
continuously
inputted so
there are
longer periods
of running. If
there is a
shortage of
any nutrients
or microbes,
more can be
placed in.
Efficient and
economical as
the fermenter
is properly
used up.
Contamination
results in
greater losses
and there may
be blocks at
inlet or outlet
pipes. There
can only be 1
process taking
place and thus
a small variety
of products.
Recombinant DNA Technology uses Enzymes
Batch and Continuous Culture:
Products may be produced in either of the 2 ways.
What is it?
Advantages
Disadvantages
Batch
Culture:
1 batch of
nutrients and
microbes are
inputted, the
process takes
place and
product is
collected and
kept separate
from other
product, the
fermenter is
empty and
sterilized and
the process is
allowed to
repeat over
and over
again.
If any
contamination
occurs, only 1
batch is
spoiled, the
culture is pure
due to
constant
sterilization
and the
fermenter can
be used in
different ways
with different
microbes
(larger variety
of product)
It may be
expensive as
the fermenter
needs to be
sterilized and
replacements
are needed
which need a
cost, thus not
being efficient.
The steps of this process are:
(1) Plasmid take from bacterial cell and cut open
using restriction endonuclease enzyme.
(2) Gene responsible for production of insulin is
taken out of a healthy human using the same
enzyme.
(3) The gene is combined to the plasmid using an
enzyme known as ligase.
(4) Recombinant plasmid placed back into the
bacterial cell.
(5) Selected bacteria are cultured in a biofermenter under optimum conditions and are
allowed to reproduce (asexually) so that more
insulin is collected.