ENZYMES

• Enzymes are protein molecules produced by living cells. Each cell contains several hundred enzymes.
They are used 'to promote a vast number of rapid chemical reactions between temperature limits
suitable for the particular organism, that is approximately 5-40 °C. In order to achieve the same
speeds of reaction outside the organism, high temperatures would be necessary, as well as marked
changes in other conditions.
• These would be lethal to a living cell, which operates in such a way as to prevent any marked change
in its normal working conditions. Thus, enzymes can be defined as biological catalysts, that is they
speed up reactions. They are vitally important because in their absence reactions in the cell would be
too slow to sustain life.
• (ATP is adenosine triphosphate, ADP is adenosine diphosphate and P1 is inorganic phosphate.) An
example of an enzyme involved in catabolism is maltase:
amylase
• starch + water ► maltose
• Commonly, a number of enzymes are used in sequence to convert
one substance into one or several products via a series of
intermediate compounds. The chain of reactions is referred to as a metabolic
pathway. Many such pathways can proceed simultaneously in the cell. The
reactions proceed in an integrated and controlled manner and this can be
attributed to the specific nature of enzymes. A single enzyme generally
will catalyse only a single reaction. Thus, enzymes serve to control the
chemical reactions that occur within cells and ensure that they proceed at an
efficient rate.
• Enzyme reactions may be either anabolic (involved in synthesis) or catabolic (involved in breakdown).
The sum total of all these reactions in a living cell or organism constitutes its metabolism. Therefore,
metabolism consists of anabolism and catabolism. An example of an enzyme involved in anabolism is
glutamine synthetase: glutamine
• glutamic acid + ammonia + ATP → glutamine + water + ADP + P1
synthetase
Catalysis and energy of activation
• Biological catalysts (that is enzymes) possess the following major
properties : all are globular proteins; they increase the rate of a
reaction without themselves being used up; their presence does not
alter the nature or properties of the end product(s) of the reaction; a
very small amount of catalyst effects the change of a large amount of
substrate; their activity varies with pH, temperature, pressure and
substrate and enzyme concentrations; the catalysed reaction is
reversible; they are specific, that is an enzyme will generally
catalyse only a single reaction.
• Consider a mixture of petrol and oxygen maintained at room
temperature. Although a reaction between the two substances is
thermodynamically possible, it does not occur unless energy is
applied to it, such as a simple spark. The energy required to make
substrates react is called activation energy. The greater the amount of
activation energy required; the slower will be the rate of reaction at a
given temperature. Enzymes, by functioning as catalysts, serve to
reduce the activation energy required for a chemical reaction to take
place. They speed up the overall rate without altering, to any great
extent, the temperature at which it occurs.
An enzyme combines with its substrate to form a short-lived
enzyme/substrate complex. Within this complex the chances of
reactions occurring are greatly enhanced. Once a reaction has
occurred, the complex breaks up into products and enzyme. The
enzyme remains unchanged at the end of the reaction and is free to
interact again with more substrate.
substrate + enzyme→ enzyme/substrate complex→ enzyme/product(s)
Mechanism of enzyme action
Detailed study has revealed that most en zymes are far larger molecules than the
substrates they act on and that only a very small portion of the enzyme, between
3-12 amino acids, comes into direct contact with the substrate in the
enzyme/substrate complex. This region is called the active site of the enzyme. It is
here that binding of the substrate or substrates occurs. The remaining amino acids,
which comprise the bulk of the enzyme, function to maintain the correct globular
shape of the molecule which, as will be explained below, is important if the active
site is to function at the maximum rate.
Enzymes are very specific, and it was suggested by Fischer in 1890 that this was
because the enzyme had a particular shape into which the substrate or substrates
fit exactly. This is often referred to as the 'lock and key' hypothesis, where the
substrate is the key whose shape is complementary to the enzyme or lock.
When an enzyme/substrate complex is formed it is 'activated' into forming the
products of the reaction. Once formed, the products no longer fit into the active
site and escape into the surrounding medium, leaving the active site free to receive
further substrate molecules.

In 1959 Koshland suggested a modification to the 'lock and key analogy. Working
from evidence that suggested that enzymes and their active sites were physically
rather more flexible structures than hitherto described, he envisaged a dynamic
interaction occurring between en zyme and substrate.
He argued that when a substrate combines with an enzyme, it induces
changes in the enzyme structure. The amino acids which constitute
the active site are moulded into a precise formation which enables the
enzyme to perform its catalytic function most effectively.
This is called the 'induced-fit' hypothesis. A suitable analogy would be
that of a hand changing the shape of a glove as the glove is put on.
Further refinements to the hypothesis have been made as details of
individual reactions became known. In some cases, for example, the
substrate molecule changes shape slightly before binding.
Factors that affect enzyme action
The effect of temperature on the rate of enzyme activity
The effect of temperature on the rate of activity of a typical enzyme. At low
temperatures, the reaction takes place only very slowly. This is because molecules
are moving relatively slowly. In other words, their kinetic energy is relatively low.
Substrate molecules will not often collide with the active site of the enzyme. As
temperature rises, the kinetic energy of the molecules increases and so the enzyme
and substrate molecules move faster. Collisions happen more frequently, so that
substrate molecules enter the active site more often. Also, when substrate and
enzyme molecules collide, they do so with more energy. This makes it easier for
bonds to be formed or broken so that the reaction can occur.
As temperature continues to increase, kinetic energy increases so the speed of
movement of the substrate and enzyme molecules also continues to increase.
However, above a certain temperature, the enzyme noo molecule vibrates so much
that some of the bonds Hq holding the enzyme molecule in its precise shape
begin to break. This is especially true for hydrogen bonds. Tods The enzyme's
active site begins to lose its shape and therefore its activity: it is said to be
denatured. At first, the substrate molecule fits less well into the active site of the
enzyme, so the rate of the reaction begins to slow down. Eventually the substrate
no longer fits at all and the reaction stops (the rate becomes zero)
The temperature at which an enzyme catalyses a reaction at the
maximum rate is called the optimum temperature. Most human
enzymes have an optimum temperature of around 40°C. By keeping our
body temperatures at about 37 °C, we ensure that enzyme-catalysed
reactions occur at close to their maximum rate.
Enzymes from other organisms may have different optimum
temperatures. Some enzymes, such as those found in bacteria which
live in hot springs (Figure 3.9), have much higher optimum
temperatures. Some plant enzymes have lower optimum temperatures,
depending on their habitat.
The effect of pH on the rate of
enzyme activity
The effect of pH on the rate of activity of a typical enzyme. Most
enzymes work fastest at a pH of somewhere around 7, that is, in fairly
neutral conditions. Some, however, have a different optimum pH. For
example, pepsin, an enzyme found in the acidic conditions of the
stomach, has an optimum pH of blood about 1.5. Pepsin is a protease,
an enzyme that catalyses the digestion of proteins.
pH is a measure of the concentration of hydrogen ions in a solution.
The lower the pH, the higher the hydrogen ion concentration. Hydrogen
ions are positively charged, so they are attracted to negatively charged
ions and repelled by positively charged ions. Hydrogen ions can
therefore interact with any charged R groups on the amino acids of
enzyme molecules. This may break the ionic bonding between the R
groups (Chapter 2), which affects the three-dimensional structure of
the enzyme molecule. The shape of the active site may change and
therefore reduce the chances of the substrate molecule fitting into it. A
pH which is different from the optimum pH can cause denaturation of
an enzyme.
When investigating pH, you can use buffer solutions (Chapter P1,
Section P1.3, Variables and making ugi measurements). Buffer solutions
each have a particular pH and maintain that pH even if the reaction
taking place would otherwise cause the pH to change. You add a
measured volume of the buffer to your reaction mixture.