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1) NATURE OF
ENZYME ACTION
DEFINITION of ENZYMES
Biological catalysts that are protein
molecules in nature.
Produced by living cells (animal,
plant & microorganism).
Absolutely essential as catalysts in
biochemical reactions.
FUNCTION of ENZYMES
To catalyze the making and breaking
of chemical bonds.
To increase the rate of reaction
without themselves undergoing
permanent chemical changes.
ENZYME REACTIONS
ENERGY OF ACTIVATION
ENERGY OF ACTIVATION
ENERGY OF ACTIVATION
In reactions occurring outside of living
organisms, simply adding heat can
provide activation energy.
Inside of living organisms, however,
another method must be used. Heat is
very harmful to the cells and proteins of
plants and animals.
ENERGY OF ACTIVATION
To overcome the activation energy required for
certain reactions to take place, living organisms
employ enzymes. Enzymes function by being
able to LOWER the activation energy needed in
specific reactions.
With a lower energy requirement, more
molecules will be able to react with each other
and the reaction can swiftly occur at
temperatures able to support life.
ENERGY OF ACTIVATION
ENZYME REACTIONS
vs
CHEMICAL REACTIONS
Highly specific - catalyze only one or small
number of chemical reactions
Their rate of reaction is usually much faster
than non-biological catalyst
The reaction conditions ( T, P, pH, and so on)
are very mild.
Enzymes are comparatively sensitive or
unstable molecules and require care in their
use.
NOMENCLATURE of
ENZYMES
*Non-descriptive name such as:
curding of milk to start cheese-making
rennin
processer
NOMENCLATURE of
ENZYMES
*Name of substrate + ase :
Substrate
Enzyme
Product
Starch
-amylase
lactose
fat
maltose
lactase
lipase
maltase
glucose + maltose +
oligosaccharides
glucose + galactose
fatty acids + glycerol
glucose
urea + H2O
urease
2NH3 + CO2
cellobiose
cellobiase
glucose
NOMENCLATURE of
ENZYMES
*Reaction which is catalyzed + ase :
Enzymes
alcohol dehydrogenase
glucose isomerase
Reaction
ethanol + NAD+
acetaldehyde + NADH2
glucose fructose
glucose oxidase
D-glucose + O2 + H2O
gluconic acid
COMMERCIAL APPLICATION
of ENZYMES
Industrial
enzymes
ENZYME KINETICS
KINETICS?????
ENZYME ACTION
LOCK AND KEY MODEL
An enzyme binds a substrate in a region called
the active site.
Only certain substrates can fit the active site.
Amino acid R groups in the active site help
substrate bind.
Enzyme-substrate complex forms.
Substrate reacts to form product.
Product is released.
ENZYME ACTION
LOCK AND KEY MODEL
ENZYME ACTION
LOCK AND KEY MODEL
ENZYME ACTION
INDUCED FIT MODEL
Enzyme structure flexible, not rigid.
Enzyme and flexible active site adjust shape to bind
substrate.
This sudden change in shape can lead to the breaking of
bonds within a single substrate molecule, forming two
new molecules. Conversely, it can also bring twosubstrate molecules close enough together for them to
bond with each other, forming one new molecule.
Increases range of substrate specificity.
Shape changes also improve catalysis during reaction.
ENZYME ACTION
INDUCED FIT MODEL
ENZYME ACTION
INDUCED FIT MODEL
E = free enzyme
S = substrate
ES = enzyme-substrate complex
P = product
2) MICHAELISMENTEN EQUATION,
LINEWEAVER BURK &
OTHER PLOTS
MICHAELIS-MENTEN
EQUATION
Leonor Michaelis (1913): noticed that at constant
[enzyme] the rate of a reaction increases with
increasing [S] until a maximal velocity (Vmax) is
achieved
This saturation effect is an important distinction
versus uncatalyzed reactions
Interpretation of data: ES complexes formed until
substrate saturation occurs at which point no more
substrate binding sites (i.e., enzyme molecules) are
available
MICHAELIS-MENTEN
EQUATION
MICHAELIS-MENTEN
EQUATION
max S
K m S
MICHAELIS-MENTEN
EQUATION
MICHAELIS-MENTEN
EQUATION
LINEWEAVER-BURK PLOT
LINEWEAVER-BURK PLOT
LINEWEAVER-BURK PLOT
3) STEADY STATE
APPROACH
ENZYME ACTIVITY
Enzymes are required in minute quantities and
enhance reaction rates by 1010 to 1020fold
When the enzyme is part of a crude preparation,
its concentration is in terms of units
Enzyme activity is the ability of an enzyme to
modify a reactant. 1 unit (U) is the enzyme
activity that converts 1 mole of reactant per
min under standard conditions.
The specific activity of an enzyme is defined as
the activity per unit of mass or U/mg protein
4) ENZYME INHIBITION:
COMPETITIVE & NONCOMPETITIVE
INHIBITION
ENZYME INHIBITION
Inhibitors
ENZYME INHIBITION
Inhibitors
Irreversible inhibitors form covalent or
very tight permanent bonds with the active
site of the enzyme and incapacitating the
enzyme. 3 classes: group-specific reagents,
substrate analogs, suicide inhibitors.
ENZYME INHIBITION
Inhibitors
Reversible inhibitors form an EI complex
that can be dissociated back to enzyme and
free inhibitor. 3 groups based on their
mechanism of action:
competitive, non-competitive and
uncompetitive.
COMPETITIVE INHIBITION
A competitive inhibitor
Has a structure similar to substrate
Occupies active site
Competes with substrate for binding to enzyme (active
site)
E + S = ES or E + I = EI . Both S and I cannot bind
enzyme at the same time
In presence of I, the equilibrium of E + S = ES is shifted
to the left causing dissociation of ES.
This can be reversed / corrected by increasing [S]
Vmax is not changed, Km is increased
NON-COMPETITIVE
INHIBITION
A noncompetitive inhibitor
Inhibitor binding site is distinct from substrate binding
site, it does not have a structure like substrate
Binds to the enzyme but not active site
Changes the shape of enzyme and active site
Substrate cannot fit altered active site
Can bind to free enzyme E and to ES
E + I = EI, ES + I = ESI or EI + S = ESI
NON-COMPETITIVE
INHIBITION
A noncompetitive inhibitor
5) TEMPERATURE &
pH EFFECT
FACTOR AFFECTING
REACTION RATE
The study of enzyme reaction rates is called
enzyme kinetics. Enzyme kinetics are affected by:
Temperature and pH: Each enzymatic reaction
has an optimum pH and optimum temperature.
Extreme temperature or pH disrupts enzyme
structure and therefore reaction rate
Substrate concentration: The reaction rate = k [P]
/ [S]. The rate can be increased by adding more
substrate, or by removing product as it is formed
FACTOR AFFECTING
ENZYME ACTION
TEMPERATURE
Little activity at low temperature
Rate increases with temperature
Most active at optimum temperatures
(usually 37C in humans)
Activity lost with denaturation at high
temperatures
FACTOR AFFECTING
ENZYME ACTION
TEMPERATURE
FACTOR AFFECTING
ENZYME ACTION
SUBSTRATE
CONCENTRATION
Increasing substrate concentration increases
the rate of reaction (enzyme concentration is
constant).
Maximum activity reached when all of enzyme
combines with substrate
FACTOR AFFECTING
ENZYME ACTION
SUBSTRATE CONCENTRATION
FACTOR AFFECTING
ENZYME ACTION
pH
FACTOR AFFECTING
ENZYME ACTION
pH
END OF CHAPTER 2