Section 1 Chemical Constituents of Life

Chapter

Enzymes
16
The enzymes speak :
“We are the catalysts of the living world !
Protein in nature, and in action specific,
rapid and accurate;
Huge in size but with small active centres;
Highly exploited for disease diagnosis in lab centres.”

E nzymes are biocatalysts – the catalysts of life.

A catalyst is defined as a substance that
increases the velocity or rate of a chemical
In the laboratory, hydrolysis of proteins by a
strong acid at 100°C takes at least a couple of
days. The same protein is fully digested by the
reaction without itself undergoing any change in enzymes in gastrointestinal tract at body
the overall process. temperature (37°C) within a couple of hours.
This remarkable difference in the chemical
The student-teacher relationship may be a
reactions taking place in the living system is
good example to understand how a catalyst
exclusively due to enzymes. The very existence
works. The students often find it difficult to learn
of life is unimaginable without the presence of
from a text-book on their own. The teacher
enzymes.
explains the subject to the students and increases
their understanding capability. It is no wonder
that certain difficult things which the students
take days together to understand, and sometimes HISTORICAL BACKGROUND
do not understand at all – are easily learnt under
the guidance of the teacher. Here, the teacher
Berzelius in 1836 coined the term catalysis
acts like a catalyst in enhancing the
(Greek : to dissolve). In 1878, Kuhne used the
understanding ability of students. A good teacher
word enzyme (Greek : in yeast) to indicate the
is always a good catalyst in students’ life!
catalysis taking place in the biological systems.
Enzymes may be defined as biocatalysts Isolation of enzyme system from cell-free extract
synthesized by living cells. They are protein in of yeast was achieved in 1883 by Buchner.
nature (exception – RNA acting as ribozyme), He named the active principle as zymase (later
colloidal and thermolabile in character, and found to contain a mixture of enzymes),
specific in their action. which could convert sugar to alcohol. In 1926,

85
86 BIOCHEMISTRY

James Sumner first achieved the isolation and enzyme reaction (type of reaction, cofactor
crystallization of the enzyme urease from jack requirement etc.)
bean and identified it as a protein.
Enzymes are sometimes considered under two
broad categories : (a) Intracellular enzymes –
They are functional within cells where they are
NOMENCLATURE AND synthesized. (b) Extracellular enzymes – These
CLASSIFICATION enzymes are active outside the cell; all the
digestive enzymes belong to this group.
In the early days, the enzymes were given The International Union of Biochemistry (IUB)
names by their discoverers in an arbitrary appointed an Enzyme Commission in 1961. This
manner. For example, the names pepsin, trypsin committee made a thorough study of the existing
and chymotrypsin convey no information about enzymes and devised some basic principles for
the function of the enzyme or the nature of the the classification and nomenclature of enzymes.
substrate on which they act. Sometimes, the Since 1964, the IUB system of enzyme
suffix-ase was added to the substrate for naming classification has been in force. Enzymes are
the enzymes e.g. lipase acts on lipids; nuclease divided into six major classes (in that order).
on nucleic acids; lactase on lactose. These are Each class on its own represents the general type
known as trivial names of the enzymes which, of reaction brought about by the enzymes of that
however, fail to give complete information of class (Table 6.1).

TABLE 6.1 Classification of enzymes

Enzyme class with examples* Reaction catalysed

1. Oxidoreductases
Alcohol dehydrogenase (alcohol : NAD+ oxidoreductase E.C. 1.1.1.1.), Oxidation o Reduction
cytochrome oxidase, L- and D-amino acid oxidases AH2 + B o A + BH2
2. Transferases
Hexokinase (ATP : D-hexose 6-phosphotransferase, E.C. 2.7.1.1.), Group transfer
transaminases, transmethylases, phosphorylase A – X + B o A + B – X
3. Hydrolases
Lipase (triacylglycerol acyl hydrolase E.C. 3.1.1.3), choline Hydrolysis
esterase, acid and alkaline phosphatases, pepsin, urease A – B + H2O o AH + BOH
4. Lyases
Aldolase (ketose 1-phosphate aldehyde lyase, E.C. 4.1.2.7), Addition o Elimination
fumarase, histidase A – B + X – Y o AX – BY
5. Isomerases
Triose phosphate isomerase (D-glyceraldehyde 3-phosphate Interconversion of isomers
ketoisomerase, E.C. 5.3.1.1), retinol isomerase, A o Ac
phosphohexose isomerase
6. Ligases
Glutamine synthetase (L-glutamate ammonia ligase, E.C. 6.3.1.2), Condensation (usually dependent on ATP)
acetyl CoA carboxylase, succinate thiokinase A+B A–B
ATP ADP + Pi

* For one enzyme in each class, systematic name along with E.C. number is given in the brackets.
Chapter 6 : ENZYMES 87

1. Oxidoreductases : Enzymes involved in made up of apoenzyme (the protein part) and a

oxidation-reduction reactions. coenzyme (non-protein organic part).
2. Transferases : Enzymes that catalyse the Holoenzyme o Apoenzyme + Coenzyme
transfer of functional groups. (active enzyme) (protein part) (non-protein part)

3. Hydrolases : Enzymes that bring about The term prosthetic group is used when the
hydrolysis of various compounds. non-protein moiety tightly (covalently) binds
with the apoenzyme. The coenzyme can be
4. Lyases : Enzymes specialised in the separated by dialysis from the enzyme while the
addition or removal of water, ammonia, CO2 etc. prosthetic group cannot be.
5. Isomerases : Enzymes involved in all the The word monomeric enzyme is used if it is
isomerization reactions. made up of a single polypeptide e.g. ribo-
6. Ligases : Enzymes catalysing the synthetic nuclease, trypsin. Some of the enzymes which
reactions (Greek : ligate—to bind) where two possess more than one polypeptide (subunit)
molecules are joined together and ATP is used. chain are known as oligomeric enzymes e.g.
lactate dehydrogenase, aspartate trans-
[The word OTHLIL (first letter in each class) carbamoylase etc. There are certain multienzyme
may be memorised to remember the six classes complexes possessing specific sites to catalyse
of enzymes in the correct order]. different reactions in a sequence. Only the native
Each class in turn is subdivided into many intact multienzyme complex is functionally active
sub-classes which are further divided. A four and not the individual units, if they are separated
digit Enzyme Commission (E.C.) number is e.g. pyruvate dehydrogenase, fatty acid synthase,
assigned to each enzyme representing the class prostaglandin synthase etc. The enzymes exhibit
(first digit), sub-class (second digit), sub-sub class all the general properties of proteins (Chapter 4).
(third digit) and the individual enzyme (fourth
digit). Each enzyme is given a specific name Genetic engineering
indicating the substrate, coenzyme (if any) and and modified enzymes
the type of the reaction catalysed by the enzyme. Recent advances in biotechnology have made
Although the IUB names for the enzymes are it possible to modify the enzymes with desirable
specific and unambiguous, they have not been characters-improved catalytic abilities, activities
accepted for general use as they are complex under unusual conditions. This approach is
and cumbersome to remember. Therefore, the required since enzymes possess enormous
trivial names, along with the E.C. numbers as potential for their use in medicine and industry.
and when needed, are commonly used and
widely accepted. Hybrid enzymes : It is possible to rearrange
genes and produce fusion proteins. e.g. a hybrid
enzyme (of glucanase and cellulase) that can
more efficiently hydrolyse barley E-glucans in
CHEMICAL NATURE AND beer manufacture.
PROPERTIES OF ENZYMES Site-directed mutagenesis : This is a
technique used to produce a specified mutation
All the enzymes are invariably proteins. In at a predetermined position in a DNA molecule.
recent years, however, a few RNA molecules The result is incorporation of a desired amino
have been shown to function as enzymes. Each acid (of one’s choice) in place of the specified
enzyme has its own tertiary structure and specific amino acid in the enzyme. By this approach, it
conformation which is very essential for its is possible to produce an enzyme with desirable
catalytic activity. The functional unit of the characteristics. e.g. tissue plasminogen activator
enzyme is known as holoenzyme which is often (used to lyse blood clots in myocardial
88 BIOCHEMISTRY

2. Concentration of substrate
Increase in the substrate concentration
gradually increases the velocity of enzyme
reaction within the limited range of substrate
levels. A rectangular hyperbola is obtained when
Enzyme velocity

velocity is plotted against the substrate

concentration (Fig.6.2). Three distinct phases of
the reaction are observed in the graph (A-linear;
B-curve; C-almost unchanged).
Order of reaction : When the velocity of the
reaction is almost proportional to the substrate
concentration (i.e. [S] is less than Km), the rate of
O the reaction is said to be first order with respect
Enzyme concentration
to substrate. When the [S] is much greater than
Fig. 6.1 : Effect of enzyme Km, the rate of reaction is independent of
concentration on enzyme velociy. substrate concentration, and the reaction is said
to be zero order.

infarction) with increased half-life. This is Enzyme kinetics and Km value

achieved by replacing asparagine (at position The enzyme (E) and substrate (S) combine
120) by glutamine. with each other to form an unstable enzyme-
substrate complex (ES) for the formation of
In recent years, it has also become possible to
product (P).
produce hybrid enzymes by rearrangement of k1 k3
genes. Another innovative approach is the E+S ES E+P
k2
production of abzymes or catalytic antibodies, Here k1, k2 and k3 represent the velocity
the antibody enzymes. constants for the respective reactions, as
indicated by arrows.
Km, the Michaelis-Menten constant (or Brig’s
FACTORS AFFECTING and Haldane’s constant), is given by the formula
ENZYME ACTIVITY k2  k3
Km
k1
The contact between the enzyme and The following equation is obtained after
substrate is the most essential pre-requisite for suitable algebraic manipulation.
enzyme activity. The important factors that
V max [S]
influence the velocity of the enzyme reaction are v= equation (1)
Km  [S]
discussed hereunder
where v = Measured velocity,
1. Concentration of enzyme
Vmax = Maximum velocity,
As the concentration of the enzyme is
S = Substrate concentration,
increased, the velocity of the reaction
proportionately increases (Fig.6.1). In fact, this Km = Michaelis – Menten constant.
property of enzyme is made use in determining Let us assume that the measured velocity (v)
1
the serum enzymes for the diagnosis of diseases. is equal to 2 Vmax. Then the equation (1) may be
By using a known volume of serum, and keeping substituted as follows
all the other factors (substrate, pH, temperature
1 V max [S]
etc.) at the optimum level, the enzyme could be V max =
assayed in the laboratory. 2 Km [S]