Enzyme

Characteristics
Catalyst
Methods
Factors
MOA
Lock-
Key
Reaction
Activation
energy
Learning Objective
Explain that enzymes are globular
proteins that catalyse metabolic
reactions.
Learning Objective
Explain the mode of action of enzyme
in terms of an active site, enzyme
substrate complex, lowering of
activation energy and enzyme
specificity.
Types Of Enzymes
exist throughout our entire body system - in
our organs, bones, blood and cells.
Grow new cells and maintain every tissue in
our body. When these enzymes are healthy,
robust and are present in adequate numbers,
they will do their job well
Metabolic
enzymes
are secreted by our various body organs - by
our salivary glands, stomach, pancreas and
small intestine.
Help in the digestion of our food.
Digestive
enzymes
METABOLISM
Catabolism
(break down)
Anabolism
(synthesis)
Break down into simpler
molecules with release
of energy
Synthesis of complex
molecules which
requires energy.
What are enzymes??
Biological catalyst
Speeds up a chemical reaction by
lowering the activation energy
Remains unchanged at the end.


What are enzymes??
Protein molecule
Globular protein -Coiled 3D
-Hydrophilic R group outside
molecule soluble
Active site Hydrophobic depression for
substrate molecule(s) to bind specifically,
form bonds with R group, catalyse
reaction

Enzyme substrate complex




Enzymes are highly specific in terms of
what they act upon and what they do.
This specificity and what the enzyme does
is made possible by the active sites.

Enzymes
Globular
protein
Biological
catalyst
Efficient &
Highly
specific
Activity is affected by
pH, temperature,
substrate
concentration and
enzyme concentration.
Activation energy & Transition State
Reactant must reach a high-
energy intermediate state called
the transition state before the
products are formed.
We need enzymes
to
lower down
the activation energy
without having to
increase the heat energy.
Enzymes reduce activation energy
Mechanism
of enzyme
action
Lock-and-key
hypothesis
Induced-fit
hypothesis

One key for one lock
One type of substrate only for one type
of enzymes (specific)
Substrate fits exactly
(complementary) to the active site of
the enzyme.
Modified version of lock-and-key hypothesis.
Active site is flexible and is not exactly complementary.
Active sites are slightly changed when the enzymes binds
with the substrates.
The active sight becomes fully complementary upon
binding.
Example: Adenylate kinase
Example: the breakdown of sucrose,
catalyzed by sucrase
Enzymatic Reaction
The close fit brings the molecules close
together and in the correct orientation for
reaction to take place.
It causes stressing and distortion of chemical
bonds of the substrates. This causes the bonds
to break and new bonds to form.
This makes it easier for the substrate to be
changed into the product and hence, lowering
the activation energy needed.
Methods of enzyme catalysis
Bring reactants together
Position reactants correctly for reaction
Provide a reaction surface (the active site)
Provide a suitable environment (hydrophobic)
Weaken bonds in the reactants
Lowering the activation energy for a reaction,
Thus dramatically increasing the rate of the
reaction.


Substrate:
sucrose bonded
together
Substrate binds to
the enzyme
forming an
enzyme-substrate
complex
3. Binding of substrate
and enzyme places
stress on the glucose
fructose bond and the
bond breaks

A non-protein component of enzymes is called the
cofactor. If the cofactor is organic, then it is
called a coenzyme.
The coenzymes make up a part of the active site,
since without the coenzyme, the enzyme will not
function.

Apoenzyme - An enzyme that requires a cofactor
-Inactive enzyme, activation of the enzyme
occurs upon binding of an organic
(Coenzyme) or inorganic cofactor.

Holoenzyme - An apoenzyme together with its cofactor.
- Complete and catalytically active.
- Contains multiple protein subunits (DNA polymerase)
- Examples: DNA polymerase and RNA polymerase
Learning Objective
Follow the time course of an enzyme-
catalysed reaction by measuring rates of
formation of products (for example catalase)
or rates of disappearance of substrate (for
example using amylase).
Understanding an enzymatic reaction
Investigating the rate at which the substrate is
converted into the product in an enzyme
controlled reaction.
amount of substrate
changed
amount of products
formed
Catalase is an enzyme found in tissues of most
living thing which catalyze the breakdown of
H
2
O

2
(hydrogen peroxide) into H
2
O and O
2
.


----------------------------------

Large volume of O
2

collected in the first
minute
As the reaction
continues, rate of
O
2
released
gradually slows
down.
Eventually the
reaction stops
because all the
substrates are
broken down
Volume of O
2

collected (cm
3
)


Time (s)
https://www.youtube.com/watch?v=w-nsa7DeXnY
Enzyme concentration (cm
3
of celery extract)
The rate of reaction in the first 30 s for each enzyme concentration.
Enzyme
concentration
Substrate
concentration
Temperature
pH
Factors that
affect the rate
of reaction
Increasing enzyme concentration increases
the rate of the reaction (if substrate is in
EXCESS/SUFFICIENT).
Enzyme
concentration
Increasing enzyme
concentration increases
the number of active sites
available to catalyze the
reaction.
If substrate in excess
Rate of reaction
Enzyme concentration
If NO substrate in excess
Rate of reaction
Enzyme concentration
Increasing substrate
concentration, maximum
plateau is reached.
Concentration of catalase
remains constant and the H
2
O

2

is the independent variable.
Substrate
concentration
Substrate concentration
R
a
t
e

o
f

r
e
a
c
t
i
o
n

V
max

Increasing temperature will
increase the rate of reaction up
to an optimum rate (highest
reaction rate).
Temperature
Increasing temperature
increases Es and Ss
kinetic energy.
E and S collide more
often per quantity of
time.
Form more E-S
complex.
Temperatures above the optimum cause increasing
enzyme molecule vibration, breaking down internal
bonds and destroying the active site.
The enzymes become denatured (lose its shape and
activity).
Temperature
Enzymes are sensitive to pH.
Operates in a narrow range on pH (either
extreme sides will denature.)

pH
How does pH affect an enzymes activity?
pH is the measure of the concentration of
hydrogen ions in a solution.
The lower the pH the higher the concentration of
hydrogen ions.
Hydrogen ions can interact with the R groups of
amino acids, affecting the way in which they
bond with each other and therefore affect their
3D structure arrangement.
A pH which is very different from the optimum
pH can cause denaturation of an enzyme.

Enzymes
Functions
Factors that
affect enzymes
activity
Mode of action
Inhibitors
Enzyme inhibition
When an inhibitor
Binds temporarily
Binds permanently
Binds to the active site
Does NOT bind to the active site
Enzyme
Inhibition
Competitive
Temporary
(active site)
Non-
competitive
Temporary
(allosteric site)
Permanent
(active site)
(allosteric)
COMPETITIVE
always for active sites
always temporary reversible

NON-COMPETITIVE
1) for active sites + permanent Irreversible

2) for other sites + permanent Irreversible
+ temporary Reversible
(not covered)
Competitive inhibitors bind reversibly to the
enzyme, preventing the binding of substrate.

On the other hand, binding of substrate
prevents binding of the inhibitor. Substrate
and inhibitor compete for the enzyme.

Competitive Reversible
Example of competitive inhibitors
C
2
H
6
O
2
Poisoning
Ethylene glycol is broken down
in the body into oxalic acid (a
deadly poison) by the enzyme,
alcohol dehydrogenase.
Alcohol (ethanol) acts as a
competitive inhibitor for alcohol
dehydrogenase.

C
2
H
6
O
2
Poisoning

Giving the patient large amounts
of alcohol will cause the ethanol
to compete with ethylene glycol
for the active site of alcohol
dehydrogenase.
Alcohol is the preferred substrate
for alcohol dehydrogenase so
when it is present, it binds with
the enzyme. After a while, the
ethylene glycol is harmlessly
excreted.
Example of competitive inhibitors
Sulphonamide
Sulphonamide is an antibacterial agent
WWII fight microbial infection
Sulphonamide similar in structure to PAB (
Substance essential for bacteria to synthesize folic
acid for its growth)
Sulphonamide act as competitive inhibitors to
PAB
DHPS
Malonate poison inhibits enzyme succinate dehydrogenase
Compete with substrate succinate which is important in
respiration
Example of competitive inhibitors -
Malonate
Binds to other site other than active site
temporarily
It is reversible.
When the inhibitors concentration
diminished by increasing the concentration of
the substrate.

Non-competitive (Reversible)
Inhibitor binds reversibly to the enzyme
Intermolecular bonds are formed
Induced fit alters the shape of the enzyme
Active site is distorted and is not recognised by the
substrate
Increasing substrate concentration reverse inhibition
Inhibitor is not similar in structure to the substrate
KCN inhibits Cytochrome C Oxidase (enzyme in
electron transport chain to produce ATP)
Reduces cell respiration
Reduced ATP synthesis
Example of Non-competitive (Reversible)
Potassium Cyanide (KCN)
http://prezi.com/gwsebj1mziks/potassium-cyanide/
**NOT COVERED IN SYLLABUS
Non competitive (irreversible) inhibitors
When inhibitor bind to the other sites than
active site permanently.
Increasing substrate concentration would not
reverse the reaction.
E.g.: heavy metal ions like mercury and silver
ions that can bind to other sites of the
enzymes.
Non competitive (irreversible) inhibitors
Inhibitor binds irreversibly to the active site permanently.
Covalent bond formed between the drug and the enzyme
Substrate is blocked from the active site
Increasing substrate concentration does not reverse
inhibition
Inhibitor likely to be similar in structure to the substrate

OH
OH
X
O
X
Covalent Bond
Irreversible inhibition
Example of Non-competitive inhibitors
(Irreversible) - Penicillin
Binds to active site
and (permanently).
It is irreversible.
Permanently blocks
the substrate.
No competition can
happen.
Example of Non-competitive inhibitors
(Irreversible) DFP Nerve gas
Nerve gas (diisopropylfluorophosphate) DFP,
combines with serine (amino acid) on the active site
of the enzyme acetylcholinesterase.
Enzyme inhibited, cannot bind to acetylcholine
(the substrate).
Acetylcholine accumulates.
Nerve impulses cannot be stopped
Prolonged muscle contraction
Enzymes with allosteric
sites often at start of
biosynthetic pathways

Enzyme is controlled by
the final product of the
pathway
Final product binds to the
allosteric site and switches
off enzyme

Inhibitor may have a
similar structure to the
final product

End product inhibitor
- Negative feedback inhibition
Allosteric sites
sites on the enzyme that bind to molecules

form weak, non-covalent bonds with these
molecules, causing a change in the
conformation of the enzyme.
End product inhibition

E.g.:
ATP (adenosine triphosphate) that act as an
allosteric inhibitor which inhibits
biochemical reaction.

When ATP level falls, the ATP will leave the
allosteric site and cellular respiration
continues to take place.
ON ACTIVE SITES OTHER SITES
TEMPORARY
PERMANENT
COMPETITIVE
REVERSIBLE
NON-COMPETITIVE
REVERSIBLE
NON-COMPETITIVE
IRREVERSIBLE
NON-COMPETITIVE
IRREVERSIBLE
ETHYLENE GLYCOL
MALONATE
END PRODUCT INHIBITION
PENICILLIN
DFP
DIGITALIS
HEAVY METALS IONS
PARATHION
Graph showing the comparative effects of the
different inhibitors on substrate concentration
When a competitive inhibitor is present, the
enzyme activity is decreased compared to
when no inhibitor is present. As the substrate
concentration is increased, the activity
eventually becomes the same.

You will notice that the optimum substrate
concentration is much higher.

With a non-competitive inhibitor, the
inhibition is not dependent on the substrate
concentration and the effect is similar to an
irreversible inhibitor.


What are the applications of the
knowledge of enzymes
in science and technology?
Give three (3) applications.