BIO150

Metabolism & Cell Division

Chapter 1

ENZYMES

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Learning Objectives

At the end of this topic, students should be able:

1. To state the general characteristics of enzymes

2. To relate between enzyme and activation energy

3. To describe the enzyme specificity based on: Key and Lock Model

Induced Fit Model

4. To identify the factors affecting enzyme activities

5. To describe enzymes inhibition

6. To classify types of enzymes

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Enzymes

Enzyme

A protein molecule serving as a biological catalyst, that speed upthe rate of chemical reaction by lowering the activation energywithout being consumed by the reaction.

Substrate / Reactant

A substrate on which an enzyme works

Active Site

The specific portion of an enzyme that binds the substrate bymeans of multiple weal interactions and that forms the pocket inwhich catalysis occur.

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General Characteristics of Enzymes

1. Made up of globular protein and coded for by DNA.

2. Catalysts because they can speed up chemical reactions

but stay unchanged at the end of the reaction.

3. Have active site where at this location the reaction

occurs.

4. Very specific, that is an enzyme can catalyze only a single

reaction.

5. Have the ability to lower the activation energy of the

reactions they catalyze.

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6. Work very efficient, where a very small amount of

enzyme can change a large amount of substrate into

products.

Eg: A single molecule of enzyme catalase can breakdown 40

million of hydrogen peroxide (H2O2) per second!

7. The catalyzing reaction of enzyme is reversible.

8. The activity of enzyme is affected by many factors. (eg.

Enzyme concentration, temperature and pH)

9. The presence of enzyme does not change the

properties of the end-product of the reactions.

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6EXTRA NOTES

The specificity of enzyme:

Enzymes are substrate specific.

Each enzyme has a unique 3D shape.

The 3D shape will recognize and bind only the specific

substrate. Enzyme active site must be 100% complementary

with the substrate.

Relationship Between Enzyme and

Activation Energy

Every chemical reaction between molecules involves both

bond breaking and bond forming.

Changing one molecule into another generally involves

contorting the starting molecule into a highly

unstable state before the reaction can proceed.

To reach the contorted state where bonds can change,

reactants molecules must absorb energy from their

surroundings.

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When the new bonds of the product molecules form,

energy is released as heat, and the molecules return

to stable shapes with lower energy then the contorted

state.

The amount of energy that reactants must absorb

before a chemical reaction will start (the energy

required to contort the reactants molecules so the bonds can

break), is known as free energy of activation or activation

energy, EA.

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Energy profile of an exergonic reaction

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The reactants must absorb

enough energy from the

surroundings to reach the

unstable transition state,

where bonds can break.

After bonds have broken,

new bonds form, releasing

energy to the surrounding.

Transition

state

Enzyme Specificity relating to Key and

Lock Model

1. Substrate enter active site.

2. Substrates are held in active site by weak interactions, such as

hydrogen bonds and ionic bonds.

3. Active site can lower EA and speed up the chemical reaction.

4. Substrates are converted to products.

5. Products are released.

6. Active site is available for new substrate molecule.10

Enzyme Specificity relating to Induced

Fit Model

1. Substrates enter active site, enzyme changes shape such that its active site enfolds the substrates (induced fit).

2. Substrates are held in active site by weak interactions, such as hydrogen bonds and ionic bonds.

3. Active site can lower EA and speed up the chemical reaction.4. Substrates are converted to products.5. Products are released.6. The enzyme returns to its original shape and active site is available for new

substrate molecule. 11

Induced Fit Model vs Key & Lock Model

Two models of enzyme reaction have been proposed.

According to the lock and key model, when the key

(substrate) fits the lock (active site), the chemical change

begins.

However, modern X-ray crystallographic and spectroscopic

methods show that in many cases the enzyme changes shape

when the substrate lands at the active site.

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Factors Affecting Enzyme Activities

A. Enzyme Concentration

The higher the concentration of enzyme, the faster the

rate of reaction as more substrate reacted.

However, as a reaction proceeds, the rate of reaction will

decrease as substrate (the limiting factor) will get used up.

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More enzyme molecules can react with more substrate

molecules, so the reaction rate increases.

B. Substrate Concentration

The higher the substrate concentration, the higher the rate of

reaction because more substrate molecules will be

colliding with enzyme molecules, so more product will

be formed.

However, after a certain concentration, any increase will have

no effect on the rate of reaction since the enzyme become

limited (limiting factor).

The enzymes become saturated, and will be working at their

maximum possible rate.

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C.Temperature

Each enzyme has an optimal temperature at which its

reaction rate is the greatest.

The rate of enzymatic reaction increases with increasing

temperature. However, above the optimal temperature, the

speed of enzymatic reaction drops sharply as the enzyme

denatured.

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D. pH

The activity of enzyme is strongly affected by changes of pH.

Different enzymes have different optimum pH values.

This is the pH value at which the bonds within them are

influenced by H+ and OH- ions in such a way that the shape

of their active site is the most complementary to the shape of

their substrate.

At the optimum pH, the rate of reaction is at an optimum.

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Negative Feedback Inhibition (End-

product Inhibition)

How does cell regulate enzymatic activity?

If the product of a series of enzymatic reactions such as

amino acid, begins to accumulate within the cell, the

product act as an allosteric inhibitor and it may

specifically inhibit the action of the first enzyme

involved in its synthesis. Thus the product begins to switch

off its own production as it accumulates.

The process is self-regulatory.

When the product being used up, its production is switched

back on again.

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Negative Feedback Inhibition

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For eg: the final product of enzyme 4 is histidine. An

increasing concentration of histidine slowly turn off the

activity of enzyme 1.

Allosteric Regulation

Another method of enzymatic control is the binding

of a regulation molecule to a protein at one site that

affects the function of the protein at a different

site.

Other than the active site, some enzymes have a

receptor site, called an allosteric site.

Substrates that affect enzyme activity by binding to the

allosteric site are called allosteric regulators.

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Allosteric Enzymes

1. The allosteric means another space, and the

characteristics of such enzyme is that they can exist in

two different forms (active and inactive).

2. The inactive form is shaped in such a way that the

substrate will not fit into the active site. For the

enzyme to work, it must be converted into the

active form (changing shape), so that the substrate

will fit into the active site.

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3. The allosteric inhibitor prevents the enzyme

from changing its shape into the active form.

The binding of allosteric inhibitor to enzyme

resembles non-competitive reversible inhibition.

4. Some allosteric regulators are activators which

result in an enzyme with a functional active site.

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Inhibition

Certain chemicals selectively inhibit the action of specific

enzymes.

Inhibition

Competitive

Non

competitive

Reversible

Irreversible

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Competitive Inhibition

a) Competitive inhibitor

mimics substrate and

compete for active

site.

Noncompetitive Inhibition

a) Noncompetitive inhibitor

alters conformation of

enzyme so active site is no

longer fully functional.

A. Competitive Inhibition

Competitive inhibitors are chemical agents that sufficiently

resemble the normal substrate.

The actual substrate thus competes for a position in the

active site. This prevents the formation of ES complex.

Eg: Succinic acid (actual substrate), malonic acid

(competitive inhibitor).

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Competitive inhibition. Malonic acid competes with succinate

for active sites of succinic dehydrogenase, an important

enzyme in the Krebs cycle

B. Non-competitive Reversible Inhibition

Occurs when an inhibitors form weak chemical bond

with the enzymes.

The inhibitors are chemical agents that bind to the site

other than active site, called an allosteric site.

It alters the shape of the enzyme, thus making the

enzyme unable to bind to the substrate.

Eg: cyanide (or potassium cyanide KCN) which combines

dehydrogenase with the cytochrome enzymes responsible

for the transfer of hydrogen atoms during cellular

respiration.

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C. Non-competitive Irreversible Inhibition

Occurs when the inhibitor first binds to the allosteric site

and then makes a covalent bond to the enzyme.

Since the inhibitor cannot fall off (bind permanently), it

changes the structure of the enzyme and makes it become

ineffective.

The enzyme thus undergoes denaturation.

Eg: Silver (Ag+), arsenic (As+)

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Enzyme Cofactors

Any non-protein molecule or ion that is required for the

proper functioning of an enzyme.

Cofactors can be permanently bound to the active site or

may bind loosely with the substrate catalysis.

It stays unchanged at the end of a reaction and can

be regenerated by a later process.

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3 Types of Cofactor

1. Inorganic ions

Known as enzyme activators.

Shape up either the enzyme or the substrate into a shape

that causes the enzyme-substrate complex to be produced.

Thus, is increasing the rate of reaction catalyzed by the

specific enzyme.

Eg: K+, Na+, Cu2+

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2. Prosthetic group

Organic non-protein substrate which is tightly

bound to the enzyme, of which it is the important part.

It is a highly active part of the enzyme molecule.

Function is to take up chemical group from the

protein part of the enzyme.

Eg: enzyme cytochrome oxidase, that plays role in

respiration, has a prosthetic group haem that contain iron.

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3. Coenzymes

Non-protein organic molecules which act as cofactors

and bind to the enzyme like prosthetic group.

Coenzymes do not bind tightly to the enzyme and do

not remain attached to the enzyme between reactions.

Most coenzymes can be classified as transfer agents

(transfer some components from one molecule to

another).

Eg: transfer electrons such as NADH, NADPH and FADH2

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Enzyme Classification

1. Oxidoreductase

Catalyze biological oxidation and reduction by the

transfer of hydrogen, oxygen, or electrons from one

molecule to another.

Eg:

Ethanal + NADH2 Ethanol + NAD

alcohol dehydrogenase

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2. Transferases

Catalyze the transfer of a chemical group from one

substrate to another.

Eg:

Glutamic acid - ketoglutaric acid

+ Aminotransferase +

Pyruvic acid Alanine

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3. Hydrolases

Catalyze the formation of 2 products from a larger

substrate molecule by hydrolytic reaction.

Eg:

Sucrose Fructose + Glucose

sucrase

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4. Lyases

Catalyze non-hydrolytic addition or removal of parts

of substrate molecules.

Eg:

Pyruvic acid Ethanol + CO2pyruvate

decarboxylase

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5. Isomerases

Catalyze internal arrangement of substrate molecule or

isomerisation

Eg:

Glucose-1-phosphate Glucose-6-phosphate

Phosphoglucomutase

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6. Ligases

Catalyze the joining together of two molecules with

simultaneous hydrolysis of ATP.

Eg:

Amino acid Amino acid-tRNA complex

+ Specific tRNA

+ ATP

Amino-acyl-tRNA

synthetase