Topic 2: Membranes, Proteins, DNA & Gene Expression

2B – Enzymes

• A catalyst is a substance that changes the rate of a reaction without changing the substances
produced.
• The catalyst is unaffected at the end of the reaction and can be used again.
• , which control the rate of the reactions that occur in cells and
organisms.
• Most of the reactions that provide cells with energy and produce new biological material would take
place very slowly at the temperature and pH of living things which is too slowly for life to exist.
• Enzymes make life possible by speeding up the chemical reactions in cells without changing the
conditions in the cytoplasm.

• For a chemical reaction to occur, the reacting molecules must have enough energy to break the
chemical bonds that hold them together.
• A simple model is that the reaction must get over an ‘energy hill’, known as the ,
before it can get started.
• Raising the temperature increases the rate of a chemical reaction by giving more molecules sufficient
energy to react.
• However, living cells could not survive the temperatures which are needed to make many cellular
reactions fast enough - and the energy demands to produce the heat would be enormous.
• Enzymes solve the problem by needed for a reaction to take place.
• The activation energy of a reaction is lowered by the following ways,
o Putting stress on the bonds within a molecule,
o or by holding molecules close together.
• This increases the likelihood of a reaction, and so lowers the energy required to begin it.

• There are two hypotheses explaining how enzymes can react.

o Lock & key hypothesis
o Induced-fit hypothesis.

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• The hypothesis states that the active site of the enzyme has a very specific shape.
• Only one substrate or type of substrate will fit the shape of the active site, and it is this that gives
each enzyme its specificity.
• The enzyme and substrate slot together to make a complex in the same way as a key fit into a lock.
• The formation of the lowers the activation energy of the reaction.
• Once the reaction is complete, the products are not the right shape to stay in the active site and the
complex breaks up.
• This releases the products and frees the enzyme for further catalytic activity.

• However, evidence from X-ray crystallography, chemical analysis of active sites and other techniques
suggests that the active site of an enzyme is not simply a rigid shape.
• The induced-fit hypothesis is generally accepted as the best current model of enzyme action.

• In this, the active site still has a distinctive shape and arrangement, but it is a flexible one.
• When a substrate molecule collides with an enzyme and if its composition is specifically correct, the
substrate enters the active site,
.
• Once the products have left the complex, the enzyme returns to its inactive, relaxed form until
another substrate molecule binds.
• This model suggests that the enzyme is like a glove and the substrate is a hand; the empty glove is
not exactly complementary in shape to the hand, but when the hand is entered it enables the glove
to mould around the hand to become completely complementary.

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1) Globular proteins.
2) The active site is a small depression on the surface of the molecule that has a specific shape because
of the way the whole large protein molecule is folded.
3) Anything affecting the shape of the protein molecule affects its ability to do the catalytic activity.
4) The three-dimensional nature of the molecule is important to the way it works.
5) A change in shape, changes the shape of the active site as well - and so the enzyme can no longer
function.
6) Enzymes change only the rate of a reaction.
7) They do not change or contribute to the end products, and they do not affect the equilibrium of
the reaction.
8) They act only as catalysts and do not modify the reaction in any other way.

1) Temperature
2) pH
3) enzyme concentration
4) substrate concentration

• Temperature affects all reactions because the number of successful collisions leading to a
reaction increases at higher temperatures.
• The effect of temperature on the rate of any reaction can be expressed as the
, .

• Between about 0 °C and 40 °C, Q10 for any reaction is 2 which means the rate of the reaction
doubles for every 10 °C rise in temperature.
• However, above 40 °C, Q10 for enzyme-catalysed reactions in human beings decreases
markedly, while Q10 for other reactions changes only slowly.
• At 60 °C the reaction stops completely in most cases as over 40 °C most proteins, including
most enzymes, start to lose their - they .
• When enzymes denature, the shape of the active site changes and so they lose their ability to
catalyse reactions.

• Some exceptions are the enzymes of thermophilic bacteria, which live in hot springs at
temperatures of up to 85 °C, can work at very high temperatures.
• They are made of temperature-resistant proteins that contain a very high density of hydrogen
bonds and disulfide bonds, which hold them together even at high temperatures.

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• pH also has a major effect on enzyme activity by
affecting the shape of protein molecules.
• Changes in pH affect the formation of the hydrogen
bonds and disulfide bonds that hold the three-
dimensional structure of the protein together which
in turn changes the shape of the active site.
• The optimum pH for an enzyme is not always the
same as the pH of its normal surroundings.
• Different enzymes work in different ranges of pH.
• This is because it is one of the ways in which cells
control the effects of their intracellular enzymes,
increasing or decreasing their activity by very small
changes in the pH.

• As the concentration of the enzyme is increased,

the enzyme activity also increases.
• Once all the substrate is bound, the reaction will no
longer speed up, since there will be nothing for the
additional enzymes to bind to.
• The substrate concentration is the limiting factor
slowing the reaction.

• Increasing substrate concentration also increases the rate of reaction to a certain point as
more enzymes will be colliding with substrate molecules.
• Once all the enzymes have bound, any substrate increase will have no effect on the rate of
reaction, as the available enzymes will be saturated and working at their maximum rate.

• The number of substrate molecules transformed per minute by a single enzyme molecule.
• Most enzymes can catalyse thousands of molecules per minute.
• Some can achieve many more: the number of molecules of hydrogen peroxide catalysed by
the enzyme catalase extracted from liver cells is 6 x 106 in 1 minute.

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• It is important to measure the initial rate of a reaction.
• Because the reaction rate will eventually slow to zero as the substrate is used up.
• So, the amount of product made per unit time should be measured at the beginning of the reaction
when the product concentration is increasing linearly.
• This value, the amount of product produced per unit time at the start of the reaction, is called the
, or V0, for that concentration.

• When V0 values are calculated for all the concentrations of interest the graph will be as below

• V0 values will increase rapidly at low substrate concentrations, then level off to a flat plateau at high
substrate concentrations.
• This plateau occurs because the enzyme is saturated, meaning that all available enzyme molecules
are already processing substrates.
• Any additional substrate molecules will simply have to wait around until another enzyme becomes
available, so the rate of reaction is limited by the concentration of enzyme.
• This maximum rate of reaction is characteristic of a particular enzyme at a particular concentration
and is known as the , or .

• The substrate concentration that gives a rate that is is called the , and is a useful
measure of how quickly reaction rate increases with substrate concentration.
• is also a measure of an for (tendency to bind to) its substrate.
• A lower Km corresponds to a higher affinity for the substrate, while a higher Km corresponds to a
lower affinity for the substrate.
• Unlike Vrnax, which depends on enzyme concentration
characterizing a given reaction.

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 • Many of the enzymes found in animals and plants work inside the cells.
• The enzymes that catalyse reactions inside cells are known as ; examples
include DNA polymerase and DNA ligase.
• Cells secrete other enzymes that work outside the cell membrane.
• The enzymes that catalyse reactions outside the cells are called . The digestive
enzymes and lysozyme, the enzyme in your tears, are good examples of these.

• Enzyme nomenclature contains the following.

o a relatively short, recommended name, which is often the name of the molecule that the
enzyme works on (the substrate) with on the end, or the substrate with an indication of
what it does (e.g., creatine kinase)
o a longer systematic name describing the type of reaction being catalysed (e.g., ATP: creatine
phosphotransferase)
o a classification number (e.g., EC 2.7.3.2).

• Some enzymes, such as urease, ribonuclease, and lipase, are known by their recommended names.
• There are still some enzymes that are known by common, but uninformative names - trypsin and
pepsin for example.
• However, the names of most enzymes give useful information about the role of the enzyme in the
cell or the body.

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• Yeast cells.

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• An intracellular enzyme is produced by a cell for action within the cell.
• An extracellular enzyme is produced by a cell and then excreted so that it acts outside the cell.

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• American chemist James B. Sumner discovered the protein nature of enzymes and developed a
general crystallization method for enzymes.
• In 1926 he became the first scientist to isolate and crystallize an enzyme, urease, found in the jack
beans.
• As this was widely believed to be impossible, his finding was ridiculed and ignored for several years,
particularly by the famed German chemist Richard Willstätter.
• In 1930 John H. Northrop was able to isolate another enzyme, proving Willstätter wrong and Sumner
right.
• In 1937 Sumner isolated and crystallized a second enzyme, catalase, found in blood.
• He shared the 1946 Nobel Prize for Chemistry with Northrop and Wendell M. Stanley.

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• Globular protein with complex 3-D shape and a critical shape to the active site
• Catalysts: speed up the rate of a reaction but are left unchanged at the end of the reaction and don’t
affect the reaction in any other way
• Enzymes are present in very small amounts.
• They are specific to one or small number of reactions of similar-shaped substrate molecules.
• rate of reaction affected by temperature and pH because these affect the shape of the enzyme and
its active site.
• rate of reaction affected by concentration of substrate only up to a certain point.