ENZYMES

Anushree Chaturvedi
 What are enzymes?
 Enzymes are highly specialized proteins
that catalyze specific biochemical reactions

 Proteins are chains of individual amino

acids
Enzymes catalyze reactions such
as these:
Starch  Glucose


-----    



Sucrose  Glucose + Fructose

  

Proteins  Amino Acids

 
Chemical reactions
 Chemical reactions need an initial input of energy =
THE ACTIVATION ENERGY
 During this part of the reaction the molecules are
said to be in a transition state.
Making reactions go faster
 Increasing the temperature makes molecules move
faster
 Biological systems are very sensitive to temperature
changes.
 Enzymes can increase the rate of reactions without
increasing the temperature.
 They do this by lowering the activation energy.
 They create a new reaction pathway ‘a short cut’
also called “ tunneling effect”
An enzyme controlled pathway

 Enzyme controlled reactions proceed 108 to 1011 times faster

than corresponding non-enzymic reactions.
Enzyme structure
 Enzymes are
proteins
 They have a
globular shape
 A complex 3-D
structure

Human pancreatic amylase

The active site
 One part of an enzyme,
the active site, is
particularly important
 The shape and the
chemical environment
inside the active site
permits a chemical
reaction to proceed
more easily
Cofactors
 An additional non-
protein molecule that is
needed by some
enzymes to help the
reaction
 Tightly bound cofactors
are called prosthetic
groups
 Cofactors that are bound
and released easily are
called coenzymes
 Many vitamins are
coenzymes Nitrogenase enzyme with Fe, Mo and ADP cofactors
 Some sources also limit the use of the term
"cofactor" to inorganic substances. An
inactive enzyme, without the cofactor is
called an apoenzyme,while the complete
enzyme with cofactor is the holoenzyme.
 Some enzymes or enzyme complexes require several
cofactors.
A good example is the multi-enzyme complex pyruvate
dehydrogenase.This enzyme complex at the junction of
glycolysis and the citric acid cycle requires five organic
cofactors and one metal ion : loosely bound thiamine
pyrophosphate (TPP), covalently bound lipoamide and flavin
adenine dinucleotide (FAD), and the co-substrates nicotinamide
adenine dinucleotide (NAD+) and coenzyme A (CoA) and a
metal ion (Mg2+).

 Organic cofactors are often vitamins or are made from vitamins

The substrate
 The substrate of an enzyme are the reactants
that are activated by the enzyme.
 Enzymes are specific to their substrates.
 The specificity is determined by the active
site.
 To explain the specificity and functioning of
enzymes different theories are proposed.
The Lock and Key Hypothesis
 Fit between the substrate and the active site of the enzyme is
exact
 Like a key fits into a lock very precisely
 The key is analogous to the enzyme and the substrate
analogous to the lock.
 Temporary structure called the enzyme-substrate complex
formed
 Products have a different shape from the substrate
 Once formed, they are released from the active site
 Leaving it free to become attached to another substrate
The Lock and Key Hypothesis

S
E
E
E

Enzyme- Enzyme may

substrate be used again
complex P

Reaction coordinate
The Lock and Key Hypothesis
 This explains enzyme specificity.
 This explains the loss of activity when
enzymes denature.
The Induced Fit Hypothesis
 Some proteins can change their shape
(conformation)
 When a substrate combines with an enzyme, it
induces a change in the enzyme’s conformation
 The active site is then moulded into a precise
conformation
 Making the chemical environment suitable for the
reaction
 The bonds of the substrate are stretched to make the
reaction easier (lowers activation energy)
The Induced Fit Hypothesis

Hexokinase (a) without (b) with glucose substrate

http://www.biochem.arizona.edu/classes/bioc462/462a/NOTES/ENZYMES/enzyme_mechanism.html

 This explains the enzymes that can react with a

range of substrates of similar types
Factors affecting enzymes:
 Temperature

 Water Content

 pH

 Chemicals

 Alteration of Substrates

 Alteration of Products
The effect of temperature
 Q10 (the temperature coefficient) = the increase in
reaction rate with a 10°C rise in temperature.
 For chemical reactions the Q10 = 2 to 3
(the rate of the reaction doubles or triples with every
10°C rise in temperature)
 Enzyme-controlled reactions follow this rule as they
are chemical reactions
 BUT at high temperatures proteins denature
 The optimum temperature for an enzyme controlled
reaction will be a balance between the Q10 and
denaturation.
The effect of temperature

Q10 Denaturation
Enzyme
activity

0 10 20 30 40 50
Temperature / °C
The effect of temperature
 For most enzymes the optimum temperature
is about 30°C
 Many are a lot lower, cold water fish will die
at 30°C because their enzymes denature
 A few bacteria have enzymes that can
withstand very high temperatures up to 100°C
 Most enzymes however are fully denatured at
70°C
The effect of pH
 Extreme pH levels will produce denaturation
 The structure of the enzyme is changed
 The active site is distorted and the substrate
molecules will no longer fit in it
 At pH values slightly different from the enzyme’s
optimum value, small changes in the charges of the
enzyme and it’s substrate molecules will occur
 This change in ionisation will affect the binding of
the substrate with the active site.
The effect of pH
Optimum pH values

Enzyme
activity Trypsin

Pepsin

1 3 5 7 9 11
pH
Substrate concentration: Non-enzymic reactions

Reaction
velocity

Substrate concentration

 The increase in velocity is proportional to the

substrate concentration
Substrate concentration: Enzymic reactions

Vmax

Reaction
velocity

Substrate concentration
 Faster reaction but it reaches a saturation point when all the
enzyme molecules are occupied.
 If you alter the concentration of the enzyme then Vmax will
change too.
Inhibitors
 Inhibitors are chemicals that reduce the rate
of enzymic reactions.
 The are usually specific and they work at low
concentrations.
 They block the enzyme but they do not
usually destroy it.
 Many drugs and poisons are inhibitors of
enzymes in the nervous system.
The effect of enzyme inhibition
 Irreversible inhibitors: Combine with the
functional groups of the amino acids in the
active site, irreversibly.
Examples: nerve gases and pesticides,
containing organophosphorus, combine with
serine residues in the enzyme acetylcholine
esterase.
The effect of enzyme inhibition
 Reversible inhibitors: These can be washed
out of the solution of enzyme by dialysis.
There are two categories.
The effect of enzyme inhibition
1. Competitive: These
compete with the
substrate molecules for E+I EI
the active site.
Reversible Enzyme inhibitor
The inhibitor’s action is reaction complex
proportional to its
concentration.
Resembles the substrate’s
structure closely.
The effect of enzyme inhibition
Succinate Fumarate + 2H++ 2e-
Succinate dehydrogenase

CH2COOH COOH CHCOOH

CH2

CH2COOH CHCOOH
COOH

Succinic acid Malonic Acid Maleic acid

The effect of enzyme inhibition
2. Non-competitive: These are not influenced by the
concentration of the substrate. It inhibits by binding
irreversibly to the enzyme but not at the active site.
(Such binding alters the shape of active site)
Examples
 Cyanide combines with the Iron in the enzymes
cytochrome oxidase.
 Heavy metals, Ag or Hg, combine with –SH groups.

These can be removed by using a chelating agent

such as EDTA.
Applications of inhibitors
 Negative feedback: end point or end product
inhibition
 Poisons snake bite, plant alkaloids and nerve
gases.
 Medicine antibiotics, sulphonamides,
sedatives and stimulants
Why are enzymes important in
the food industry?
 Added or used to cause particular
reaction
 Advantages
 Natural, Nontoxic
 Catalyze specific reactions
 Active under mild conditions
 Active at low concentrations
 Can control rate of reaction
 Can be inactivated
Enzymes as Industrial Products
 Microorganisms are ideal for the large-scale
production of enzymes. Many enzymes are
used in the laundry industry to remove stains
from clothing, and thermo stable and alkali
stable enzymes have many advantages in
these markets.
 Certain enzymes are produced in large
amounts by some organisms, and instead of
being held within the cell, they are excreted into
the medium.

 These extracellular enzymes, called

exoenzymes, can digest insoluble polymers such
as cellulose, protein, and starch. The products of
digestion are then transported into the cell where
they are used as nutrients for growth.
 The term extremozyme has been coined to describe
enzymes that function at some environmental extreme,
such as high temperatures or low pH. Enzymes from
extremophiles are desirable for biocatalysis under
extreme conditions.
 For some biocatalytic processes, it is
desirable to fix soluble enzymes onto a solid
surface. These are called immobilized
enzymes.