BIOL1: Enzymes (8)
3.1.2 The digestive system provides an
interface with the environment. Digestion
involves enzymic hydrolysis producing
smaller molecules that can be absorbed and
assimilated.
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Enzymes as catalysts lowering activation
energy through the formation of enzyme
substrate complexes.
The lock and key and induced fit models of
enzyme action.
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The properties of enzymes relating to their
tertiary structure.
Description and explanation of the effects of
temperature, competitive and noncompetitive inhibitors, pH and substrate
concentration.
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To take (something) in through.
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Physiology The conversion of nutrients into
living tissue; constructive metabolism.
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Noun: A substance that increases the rate of
a chemical reaction without itself undergoing
any permanent chemical change.
Or...........................
 A person or thing that precipitates an event.
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The material or substance on which an
enzyme acts.
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To bring about or stimulate the occurrence
of; cause.
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use the lock and key model to explain the
properties of enzymes. They should also
recognise its limitations and be able to
explain why the induced fit model provides a
better explanation of specific enzyme
properties.
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Enzymes are globular protein molecules,
generally soluble in water.
They have a 3D shape or tertiary structure.
It has hydrophobic amino acid R-groups in
the centre of the “ball” and hydrophilic amino
acid R-groups around the outside of the ball.
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The formation and breakage of glycosidic
bonds, ester bonds and peptide bonds need at
least 1 enzyme for each.
Process of protein synthesis, digestion,
respiration and photosynthesis require a
number of different enzymes.
It has been estimated that one cell may contain
over 1000 different enzymes.
Most enzymes are made up of 100s of amino
acids.
1. What do ester bonds hold together?
2. What do glycosidic bonds hold together?
3. What do peptide bonds hold together?
1. lipids/Fats. (3 fatty acids + glycerol =
triglyceride)
2. Monosaccharides (2 monosaccharides
bonded together = disaccharide.)
3. Amino acids. (Bonded together to form a
polypeptide)
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Enzymes that act inside cells = intracellular
enzymes.
E.g. - Hydrolases. These are found in lysosomes
and hydrolyse (break down) substances cell has
taken in by phagocytosis.
- ATPases found inside mitochondria. Involved in
synthesis of ATP in aerobic respiration.
Enzymes that act outside cells = extracellular
enzymes.
E.g. Digestive enzymes in the alimentary canal.
E.g. amylase which hydrolyses starch to maltose.
Enzymes have a precise 3D shape (= tertiary
structure).
Have hydrophilic R groups (side chains), on
outside of molecules, makes em soluble in water.
Often name of enzyme ends in ase (e.g. ATPase
or protease).
4. Explain why enzymes are so specific in the
reactions that they catalyse.
5. Suggest why all enzymes are protein
molecules.
6. The name given to enzymes that catalyse
the breakage of peptide bonds is proteases.
What name would be given to the enzymes
that catalyse the breakage of:
(a) Glycosidic bonds
(b) Ester bonds
4. The catalytic part of the enzyme is the active
site. This has a very specific shape. So only
one or a few molecules can fit into it.
5. Only proteins can produce such a wide
variety of shapes. This is because they are
made up from 20 different amino acid sub
units.
6. (a) carbohydrases
(b) Lipases
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Heterotrophs obtain their nutrients, (not
food), by consuming other organisms. They
need to break down the body of the organism
they are consuming, digest it, and then
absorb it.
Digestion = breaking larger units into
smaller, monomer units. So need to break the
bonds. Like peptide bonds, ester bonds, and
glycosidic bonds. These are catalysed by
different types of enzymes.
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Some organisms secrete or release enzymes
outside their body, onto the food to digest
them, then absorb the smaller monomer
units.
Others have an internal digestive system.
Food taken into body, enzymes mix with it,
digests it into monomer units and then it is
absorbed. Many of these enzymes are also
extracellular (= action takes place outside
cells) as they are released from the cells that
produce them, onto the food in the digestive
system spaces, (like the lumen of your gut).
Also get intracellular = enzymes found in the
cytoplasm of the cells or attached to the
membranes. Here their action takes place
inside cells.
3. Enzymes and protection
Many organisms use
enzymes as a
defence
mechanism.
E.g. white blood cells
called phagocytes
take in and digest
bacteria using
lysosomal enzymes
7. Suggest the advantages of having an internal
digestive system compared with secreting
enzymes outside the organism.
8. Explain why many white blood cells involved
in phagocytosis contain a high concentration
of enzymes.
9. Suggest why all organisms, no matter what
sort of environment they live in, have
enzymes in their cells.
7. Enzymes are not lost and can be recycled
internal system can be regulated to provide
the optimum conditions for the enzyme.
8. Their job is to take in and destroy foreign
organisms and debris. The destruction is
caused by digestive enzymes in the
lysosomes of these cells.
9. Enzymes regulate metabolic processes by
catalysing reactions at a rate appropriate to
the organism.
Enzymes are globular proteins, with a specific
3D shape, resulting from the sequence of
their amino acids. They are large but only a
small region is functional = active site.
Usually a cleft or depression, (dip), where
another molecule (or substrate) can bind.
Substrate is sometimes called the reactants,
(the starting materials). Random movements
bring the substrate into contact with enzyme.
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Substrate fits perfectly into active site and is
temporarily bound to some of the R groups of
the enzyme’s amino acids. The substrate
molecules move around randomly and collide
with the active site of the enzyme.
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When the molecules combine enter a
transition state – here bonds in molecules
become strained. The molecules are
activated. In this state they are more likely to
have bonds broken or new ones formed.
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One enzyme – one substrate. As only one
substrate fits into that enzyme, like a lock
and key = the lock-and-key hypothesis.
Remember the substrate does not have the
same shape as the active site. It has a
complementary shape to the active site.
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In most enzymes when substrate fits into active
site, the shape of whole enzyme changes shape,
so that it fits around the substrate more closely
to give the enzyme-substrate complex. The
enzyme can hold the substrate in position for
reaction to occur = the induced-fit hypothesis.
By altering its shape the enzyme puts a stain on
the substrate molecule, which distorts a
particular bond on the substrate and so lowers it
activation energy needed to break this bond.
Once the products are released from the
enzyme it returns to its original shape, ready
to be used again.
Whether by simple lock and key or induced fit
the enzyme is specific to this substrate.
10. Explain why changing the shape of one of
the amino acids that make up the active site,
could prevent the enzyme from functioning.
11. Why might changing certain amino acids
that are not part of the active site, also
prevent the enzyme from functioning?
10. The changed amino acid may not bind to
the substrate, which will not then be
positioned correctly, if at all in the active site.
11. The changed amino acid may be one that
forms hydrogen bonds with other amino
acids. These bonds will then not form and so
affect the tertiary structure of the enzyme.
The shape will change, including the active
site, so substrate will not fit.
Enzyme may
catalyse a
reaction
where:
- Substrate is
split into 2 or
more
molecules
(Catabolic)
Or
- 2 or more
molecules
are joined
together
(Anabolic).
(Amino acid
+ amino
acid =
dipeptides)
Interactions between the R groups of the
enzyme and the atoms of the substrate can
break or encourage formation of bonds with
the substrate molecule. So 2 or more
products are formed.
When reaction complete, product(s) leave
active site.
Enzyme unchanged, so ready to be used again.
Pretty quick reactions. E.g. enzyme catalase
can bind with hydrogen peroxide ( ‘a super
oxide’), split them into water and oxygen,
and release products at rate of 107 molecules
per second.
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Some slower. Enzyme rubisco involved in
photosynthesis can only deal with 3
molecules per second.
Rubisco is the most abundant enzyme on the
planet – why?
Most of the time the substrate won’t convert
into its product without extra energy =
activation energy (EA). = the initial input of
energy to start the reaction. Once started
reaction continues of its own accord. The role
of the enzymes is to lower this activation
energy.
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Can provide the equivalent of extra heat to a
reaction as they increase the energy of the
reactants.
Mammals do this by having a constant body
temp of 370.
This temp wouldn’t provide enough activation
energy for many substrates to work (without
enzymes) and if it goes above 400 then can
permanently damage proteins.
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Enzymes are the answer as they decrease
activation energy. Do this by holding
substrate so molecules can react more easily.
So reactions with enzymes take place at lower
temps than would do without them.
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Catabolism = substances break down and
release energy.
Anabolism = simple molecules are built up
into more complex molecules and use
energy.
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Reactions that liberate more energy than they
use = exergonic.
Reactions that take in more energy than they
liberate = endergonic.
1 – Kinetic energy = matter that is moving and
performing work has kinetic energy.
2 – Potential energy = matter that is not
performing work but has the ability to do so
is potential energy.
12. Describe how the lock-and-key hypothesis
of enzyme action differs from the induced fit
hypothesis.
13. Explain how enzymes reduce the activation
energy of a reaction.
14. Explain why the reduction in activation
energy provided by enzymes is essential to
living organisms.
12. Induced fit = enzyme changes shape to
hold the substrate in active site.
Lock-and-key = no shape change to enzyme.
13. Enzyme holds the substrate so reaction
occurs more easily and requires less
activation energy, than the reaction occurring
without an enzyme.
14. Most reactions don’t take place at a
sufficient rate to sustain life without
enzymes. They need to be faster.
Catalase is found in tissues of most living things
and catalyses breakdown of hydrogen peroxide
into water and oxygen.
Catalase
Hydrogen peroxide → Oxygen + Water
Can do this practically. As reaction happens can
collect oxygen. Reaction happens quickly at first,
and then reaction gets slower and slower until it
stops.
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Rate of reaction depends on how many enzyme
molecules there are and the speed they combine
with substrate. At first loads and loads of
available enzyme molecules for substrate to bind
with. So at beginning of reaction often number of
enzyme molecules that limits rate.
As substrate is converted into product there is
less and less substrate left. Enzymes molecules
are left waiting for substrate. Reaction gets
slower and slower then stops. This is where the
graph flattens out.
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So reaction is always fastest at the beginning
= called initial rate of reaction.
Can measure this by calculating the slope of a
tangent to the curve, as close to time 0 as
possible.
Or read off the graph the amount of O2 given
off in the first 30 seconds.
15. Why is it better to calculate the initial rate
of reaction from a curve, rather than simply
measuring how much oxygen is given off in
the first 30 seconds?
15. Risk of inaccuracy in a single measurement
at 30 seconds.
Shape of curve gives more accurate value as is
based on many readings taken over a given
period of time rather than just 1.
To compare look at the beginning of reaction
as once reaction started amount of substrate
begins to vary as substrate is converted to
product at different rates.
Only at beginning that is def that the
differences in reaction rate are caused only by
difference in enzyme conc.
If double number of enzymes double number
of active sites, as long as is plenty of
substrate.
Easy to measure rate if one of the products if it
is a gas. Not always easy though.
E.g. looking rate at which amylase breaks down
the substrate starch to the product maltose.
Both are colourless.
Could measure rate starch is used up. Take
samples at known times and test with iodine.
Could use a colorimeter can measure intensity
of blue-black colour.
Can plot graph of remaining starch against
time. Can then calculate initial reaction rate
as before.
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With this practical it is easier if you mix
starch, iodine in potassium iodide solution
and amylase and take regular readings of the
colour of the mixture in one tube in a
colorimeter. Not ideal though as the iodine
interferes with the reaction and slows it
down.
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16. Sketch the curve you would obtain if the
amount of starch remaining was plotted
against time.
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Graph below shows results of prac where
conc of catalase, (enzyme), was kept constant
and conc of hydrogen peroxide, (the
substrate), was varied. Curve shows oxygen
released against time. Initial rate was
calculated for first 30 seconds.
Initial rates of reaction were plotted against
substrate conc.
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The graph shows as substrate conc increases
so does the initial rate of reaction. This is cos
there are more substrate molecules around.
Comes a point where every enzyme active site
is occupied and enzyme can’t work any
faster. The enzyme is working at its
maximum possible rate, known as Vmax.
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At a point in the reaction, (shown on the
graph where the line levels off, known as a
plateau), either the enzyme conc or the
substrate conc prevent any further increase in
reaction rate. They are limiting the reaction =
limiting factors. If the conc of the limiting
factor is increased then the reaction rate
increases.
17. Sketch the shape that the graph above
would have if excess hydrogen peroxide was
not available.
18. Suggest why enzymes are usually
maintained at low concentration in cells.
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18. Enzymes
catalyse reaction
quickly and can be
reused.
Graph below shows how the rate varies with
temperature.
At low temps – slower reaction. This is because
molecules have less kinetic energy and so
move around slower, so collide less often.
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Higher temp – faster rate of reaction. This is
due to the molecules having a higher natural
kinetic energy and so they vibrate more and
move around faster, so collide faster rate, so
combining at a faster rate. So when they do
collide at higher temp, they have more energy
and so have sufficient activation energy to
react. It means it is easier for bonds to be
broken so reaction can occur.
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Once temp reached a certain point vibrations
of enzyme are so great that as well as making
them collide quickly, some of the weaker
bonds, (like hydrogen and ionic bonds),
which give it it’s precise shape break. Tertiary
structure held less and less in shape. Rate of
reaction decreases. This happens at approx
45o in humans. Now the substrate still fits
into the active site, but not as well.
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If enough bonds are broken then tertiary
structure is unravelled and enzyme stops
working. Enzyme loses shape = denatured.
(At approx 650). The tertiary structure is
damaged then it is not reversible. It is too
damaged to rebuild. However if only some
bonds affected and shape changed to slow
reaction but tertiary structure is still intact,
then it is often reversible.
Primary structure is not affected.
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In the human body, above 400 substrate
molecules fit less well into active site of
enzyme and so the rate slows down. As temp
goes higher substrate no longer fits so no
reactions.
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Temperature that enzyme catalyse a reaction
at its maximum rate = optimum temperature.
(For humans = 400. Our body temp is 370. It
would be dangerous to maintain a body temp
of 400, cos even a slight rise in temp would
begin to denature enzymes. So you can have
a temperature rise when you are ill and your
enzymes stay intact.).
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Mammals and birds are endothermic = able
to maintain their internal body temp
independently of the environment. Is a cost
though = they require greater amount of
food than similar sized reptiles.
Other organisms have different optimum
temps. Some bacteria that live in hot springs
have very high optimum temps – useful in
various industrial applications. Some plants
have very low ones.
19. How would you carry out an experiment to
determine the effect of temperature on the rate
of breakdown of hydrogen peroxide by catalase?
20. Explain why increased kinetic energy increases
the rate of reaction in an enzyme controlled
reaction.
21. What type of bonds would you expect to find in
greater numbers holding the tertiary structure of
heat resistant enzymes, compared with more
heat sensitive enzymes?
22. Suggest why the normal body temp of
mammals is slightly below the optimum temp of
most of the enzymes that occur in the organism.
19. Have several catalase – hydrogen peroxide
reactions going, at different temperatures. All
other factors must remain constant, including the
volume & concentration of hydrogen peroxide,
(substrate) and the catalase, (enzyme) solutions.
Measure the volume of oxygen given off over time.
20. Increased kinetic energy = increased collisions
between enzymes and substrates and so
increased rate of reactions.
21. Disulphide bonds.
22. If normal body temp was too near to the
optimum temp for the enzymes then if
organism got a fever or exercised, so heat is
generated, then enzymes may be denatured.
A higher body temp as well as denaturing
enzymes also would require more energy.
pH = a measure of the hydrogen ion
concentration. The higher the conc – the lower
the pH, (the more acidic it is).
Tertiary structure of enzyme held together by
hydrogen and ionic bonds. These bonds are due
to the attraction between oppositely charged
groups on the amino acids that make up the
enzyme.
Hydrogen ions, (because of their charge), interfere
with hydrogen and ionic bonds, so alters tertiary
structure and so changes the active site.
Minor changes in pH only affect a few
hydrogen and ionic bonds and these can
reform if the pH returns to its optimum. This
will reduce enzyme activity. Has to be an
extreme pH to denature enzyme.
All enzymes have an optimum pH, (where they
work fastest). For most this is around pH 7.
Not all though. Protease, (each enzyme in
your stomach), works best at pH 2/3. This is
because you have acid in your stomach to kill
bacteria that you may have ingested.
23. Individual hydrogen bonds are fairly weak. How
can such weak bonds be responsible for holding
the tertiary structure of enzymes in place?
24. Enzymes produced by microorganisms are
responsible for spoiling food. Using this fact and
your knowledge of enzymes, suggest a reason
why:
(a) Food is heated to a high temperature before
canning.
(b) Some foods, such as onions, are preserved in
vinegar.
23. There are many hydrogen bonds. One is
weak but together they are strong.
24a. High temps denature enzymes in
microorganisms, so they cannot spoil the
food.
b. vinegar is acidic, so has low pH. The low pH
denatures the enzymes in the
microorganisms and so preserves the food.
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Inhibitor = a substance that slows down or
stops an enzyme controlled reaction.
Other substances can have the same shape as
the substrate and so can fit into the active
site of the enzyme. This inhibits the enzyme.
Some substances fit onto another part of the
enzyme and indirectly cause a shape change
n the active site of the enzyme. Most
inhibitors affect only one enzyme, others
affect many.
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Reversible Vs irreversible
Some inhibitors can be temporary inhibitors
= only binds briefly to active site. In this case
if there are loads more substrate molecules
than inhibitors, then the substrate can still
bind and reaction rate hardly affected.
But if concentration of inhibitor rises or
substrate falls then more likely inhibitor will
collide with active site of enzyme rather than
substrate and so reaction rate slows.
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Inhibitor can remain in active site
permanently. = irreversible inhibition. Even if
more substrate added the inhibitor can’t be
displaced.
(1) If the inhibitor has the same shape as the
substrate and can fit into the active site of the
enzyme = competitive inhibition. They form
an enzyme-inhibitor complex. No product
formation cos enzyme does not catalyse a
reaction.
It is normally reversible. Can be reversed by
increasing concentration of substrate.
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(2) Sometimes a molecule can bind to another
part of the enzyme, rather than the active
site. This changes the shape of the active site
of the enzyme, by distorting the tertiary
structure of the enzyme.
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Substrate can’t bind, however much of it
there is. = non-competitive inhibition. It can
be reversible inhibition but more normally is
irreversible inhibition. Depending on whether
the inhibitor bonds briefly or permanently
with the enzyme.
Allosteric = refers to when the enzymes’
shape is altered.
When an inhibitor binds with the enzyme on a
site other than its active site it changes the
shape of the active site, so the substrate can’t
bind to it = Non-competitive inhibition.
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Whether the inhibition is
reversible or nonreversible, whilst the
inhibitor is bound to the
enzyme the enzyme is said
to be denatured.
E.g. Digitalis is a substance
extracted from the plant
foxglove. It is a noncompetitive inhibitor. It
binds with the enzyme
ATPase and results in an
increase in the contraction
of the heart muscle.
25. Suggest a method of determining
whether the inhibition of an enzymecontrolled reaction is competitive or noncompetitive.
26. Competitive inhibitor molecules can be
much larger than the substrate molecules
they compete with. Suggest how this is
possible.
25. Carry out the reaction with a range of
substrates concentrations. If the rate of
reactions increases up to the same as that
given without inhibitor present, then the
inhibitor is a competitive inhibitor.
26. The competitive inhibitor may be a large
molecule with a small part the same shape
as the substrate (and the active site of the
enzyme).
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Some poisons
inhibit the enzyme
but other over
activate the
enzyme.
Inhibitors that seriously
disrupt enzyme
controlled reactions can
act as metabolic poisons.
They prevent vital
chemical reactions taking
place.
E.g. 1. Poison found in the
deadly death cap
mushroom called alphaamanitin, inhibits
enzymes that catalyse
the production of RNA
from DNA. Cells can’t
make proteins and die.
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E.g. 2. Potassium cyanide inhibits cell
respiration. It is a non-competitive
inhibitor for the vital respiratory enzyme
cytochrome oxidase, found in
mitochondria. ATP can’t be made. The
organism can only respire anaerobically
which leads to lactic acid build up in the
blood. Only need 100-200mg for an adult
to lose consciousness, in 10s. If untreated
go into a coma in 45mins and dead after
2hrs.
Found in antifreeze. Not poisonous but when broken
down in liver by enzyme alcohol dehydrogenase,
the breakdown product called oxalic acid is very
toxic and can cause death.
Remedy = give massive dose of ethanol, (alcohol).
Leads to severe (but less likely to be fatal), alcohol
intoxication. The ethanol acts as a competitive
inhibitor of alcohol dehydrogenase. This reduces
rate of production of oxalic acid, allowing ethylene
glycol to be excreted harmlessly.
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The antibiotic penicillin works as an inhibitor.
It permanently occupies the active site of an
enzyme which helps synthesis bacterial cell
walls. Prevents new bacterial cells being
produced.
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Snake venom is a
mixture of toxins
and different
enzymes.
Phosphodiesterases
are in most
venoms.
They cause a fall in
prey’s blood
pressure and cause
heart failure.
27. Many antibiotics are chemicals that fungi
produce and release into their environment.
What is the advantage to a fungus of
producing and releasing antibiotics?
28. Suggest why proteases inhibitors can
inhibit viral proteases, but do not affect the
human protease enzymes in the cell.
27. The fungi release the antibiotics to destroy
other organisms, (bacteria), so that the other
organisms cannot take up its food supply.
(Defence mechanism).
28. Protease enzymes of viruses differ in shape
to that of the protease in humans. Protease
inhibitors are specific.
Many enzymes need the presence of another nonprotein substance to work. These substances =
coenzymes, or cofactors.
No real difference between the 2 but some use
cofactor to mean simple molecule inorganic ion
and term coenzyme for bigger molecules.
Both work by binding briefly with enzyme.
Sometimes this alters their shape so can bind
more effectively with substrate. Sometimes it
helps the enzyme to transfer a particular group
of atoms from one group to another.
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Small, inorganic non-protein molecules. Bind for a
short time to the active site, either just before or at
same time as substrate. In many reactions the
coenzymes take place in the reaction and like
substrates are changed during it. Unlike substrates
though, they can be recycled back again though.
Many cofactors are made from vitamins. E.g. vitamin
B3 (nicotinamide) is important role in breaking down
fats and carbohydrates. Vitamin B3 used to make a
coenzyme required for the enzyme pyruvate
dehydrogenase to function properly. This enzyme is
required in respiration.
Deficiency in B3 leads to disease known as pellagra.
This causes diarrhoea, dermatitis, dementia and
death eventually.
- = a coenzyme which is permanent part of an
enzyme. They contribute to final 3D shape of
enzyme. E.g. the enzyme carbonic anhydrase
contains zinc based prosthetic groups.
Enzyme essential component of red blood
cells. Involved in catalysing the combining of
carbon dioxide and water to form carbonic
acid. (Which is how CO2 transported in
blood).
small inorganic ions. May combine with enzyme or
substrate. The binding makes the enzyme-substrate
complex form more easily cos it affects the charge
distribution and sometimes the shape of the
enzyme-substrate complex.
E.g. (cofactor) chloride ions bind to salivary amylase,
changes its shape a bit and help starch to bind to if
more efficiently. Starch is then broken down into
maltose.
E.g.2 (cofactor) calcium ions essential for enzyme
thrombin, which catalyses change of soluble,
globular protein fibrinogen into the insoluble,
fibrous protein fibrin, used in blood clotting.
E.g. 3 (coenzyme) = coenzyme A. needed in many
metabolic pathways including aerobic respiration.
29 Suggest why the recommended daily dietary
allowance for nicotinamide (18mg), is very
low.
30. Name the prosthetic group found in
haemoglobin.
31. Suggest where in the cell the addition of
prosthetic groups to the enzyme molecules
take place.
31. It is a coenzyme and is reusable.
32. The haem group.
33. The Golgi apparatus.
They are specific so they do not produce a
range of unwanted products hence they have
a broad use in industry:
 28% detergents
 35% food processing
 23% beverages
 14% others