Factors Affecting Enzyme Activity and Kinetics Experiments

 
2801 June 05
 
 
2801 June 05
 
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Factors Affecting
Enzyme Activity
 
Factors Affecting Enzyme
Activity
 
O
Concentration of the substrate
O
Concentration of the enzyme
O
Temperature
O
pH
O
Presence of inhibitors
Simulating Enzyme Kinetics
O
Equipment
O
Popping beads
O
Trays
O
Stopwatches
O
Work in groups of three
A is the Enzyme
B passes the beads to the enzyme
    at a set rate (collision rate)
C manages the stop clock.
 
Results
Increasing the substrate
concentration
 
O
As concentration of substrate increases, collisions
between enzyme and substrate molecules occur
more often.
O
More enzyme-substrate complexes form
, so more
product is formed.
O
The reaction rate increases
Page 132-135
The effect of substrate concentration on the rate of an enzyme
controlled reaction
Rate of
reaction
(au)
Substrate conc (mol/dm³)
 
Why has the rate
levelled out?
What is limiting any
further increase in the
rate of reaction ?
 
Here the increase in
rate and substrate conc
are proportional
 
Exam question.
Describe the relationship between the substrate concentration
and the rate of the reaction.
 
Look for overall trends
 
Refer to points on the graph
 
Do not generalise
 |                |                  |                 |                  |
1                2                  3                 4                  5
 
7   --
6   --
5   --
4   --
3   --
2   --
1   --
Increasing the enzyme concentration
 
O
As the enzyme concentration increases, 
more active
sites become available.
O
More enzyme-substrate complexes form (and so more
products are produced). The rate of reaction increases.
 
1.
Sketch a graph to show the relationship between enzyme
concentration and the rate of reaction.
2.
What happens if the substrate concentration increases further?
3.
Draw another line to show what the reaction would look like if
there was double the substrate concentration
Enzymes and temperature
 
O
Kinetic energy and collision theory
O
Heat, vibration, breaking bonds and denaturation
Give a definition of denaturation
Describe how heating can lead to denaturation in enzyme
structure
T
e
x
t
b
o
o
k
 
p
1
2
8
-
9
 
Optimum temperatures
 
 
Refer to the graph to explain the effect of temperature on enzymes.
Include:
 
Kinetic energy, Collision theory , enzyme/substrate complexes,
denaturation
 
T
e
x
t
b
o
o
k
 
p
1
2
8
-
9
 
Precise terminology
 
Formation of enzyme substrate complexes
 
Increase in the kinetic energy of the substrate and enzyme
molecules
 
High temperatures disrupt the tertiary structure of the
enzyme by breaking the tertiary bonds
Enzymes and pH
 
O
What is pH?
O
pH and bonds
 
(interfere with hydrogen bonds
and ionic bonds-tertiary
structure affected)
T
e
x
t
b
o
o
k
 
p
1
3
0
-
1
3
1
pH and active site
T
e
x
t
b
o
o
k
 
p
1
3
0
-
1
3
1
Importance in terms
of induced-fit
hypothesis
Increasing H ions,
alters charges
around the active
site
 
Optimum pH
 
O
Varies depending on
enzyme (e.g. where
found)
O
At optimum,
concentration of H ions
in solution gives the
tertiary structure the best
overall shape.
O
Range is generally fairly
narrow
Buffer solutions at set pH values are used to carry out enzyme
reactions. The rate can then be calculated and the optimum
found.
Denaturation will only occur in extreme changes of pH away
from the optimum
. 
The further from the optimum the faster the
rate of denaturation will be.
T
e
x
t
b
o
o
k
 
p
1
3
0
-
1
3
1
Inhibitors
 
O
Competitive
 
Similar in shape to substrate
O
Non competitive
 
Do not compete with substrate molecules for a place in
the active site
O
Permanent
 
Most competitive inhibitors are not permanent  (action
is reversible).
 
Many non competitive inhibitors bind permanently to
enzyme molecules
An enzyme inhibitor is any substance or
molecule that slows down the rate of an
enzyme-controlled reaction by affecting the
enzyme molecule in some way
.
 
Enzyme inhibitors
 
Irreversible
 inhibition
 
O
The inhibitor binds permanently with the enzyme and
changes the shape of the active site preventing
substrate binding.
O
e.g. heavy metal ions – mercury, lead
Normal enzyme
substrate binding:
 
E
 
S
 
Substrate no longer fits into
active site and effect is
irreversible
Reversible
 inhibition
O
Competitive
O
Non-competitive
AS Guru enzymes
 
If inhibitor occupies the active site
and substrate can no longer bind
 
Increasing concentration of substrate
reduces effect of inhibitor
 
Inhibitor binds at another site
(
allosteric
 site) on the enzyme but
changes the shape of the active
site
 
Substrate cannot bind, even if its
concentration is increased
 
Effect is however reversible – if
inhibitor is removed shape of active
site is restored
 
e.g. Leeches produce hirudin
which inhibits the production of
a blood clotting protein
 
e.g. cyanide inhibits an enzyme
involved in respiration
Substrate concentration
Substrate concentration
 
With a competitive inhibitor
 
With  a non competitive inhibitor
Rate
Of
reaction
Rate
Of
reaction
Decide which graph represents the effect on the rate of an enzyme
controlled reaction when exposed to:
a competitive inhibitor
a non competitive inhibitor.
 Each reaction is carried out with a fixed quantity of inhibitor.
          Normal
 
rate
          Inhibited rate
End-product inhibition
O
The product of the reaction which is being catalysed
acts an inhibitor of the enzyme involved
 
products
 
substrate
1. Substrate
binds at active
site
2. Reaction is catalysed
3. Product inhibits
enzyme (usually non-
competitive)
True or false?
O
An enzyme is a biological catalyst
O
An enzyme will work faster at 60
°
C
O
An enzyme will be denatured at 100 
°
C
O
Enzymes break things down
O
Enzymes are denatured if chilled, which is why
a fridge preserves food
O
There will be as much enzyme left at the end of
a reaction, as there was at the beginning
O
Blank-ase breaks down or interacts with blank
 
True
Well
Well
False
False
 
True
 
True
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Explore the factors influencing enzyme activity, such as substrate and enzyme concentration, temperature, pH, and inhibitors. Learn how to simulate enzyme kinetics using equipment like popping beads and stopwatches. Analyze results to understand the impact of substrate concentration on reaction rates. Discover the relationship between substrate concentration and reaction rate through graphs and exam questions.


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  1. 2801 June 05

  2. 2801 June 05

  3. Factors Affecting Enzyme Activity

  4. Factors Affecting Enzyme Activity O Concentration of the substrate O Concentration of the enzyme O Temperature O pH O Presence of inhibitors

  5. Simulating Enzyme Kinetics O Equipment O Popping beads O Trays O Stopwatches O Work in groups of three A is the Enzyme B passes the beads to the enzyme at a set rate (collision rate) C manages the stop clock.

  6. Results Time Time interval interval between between passing to passing to enzyme enzyme (s) (s) 10 10 Substrate Substrate Concentration Concentration (Beads (Beads passed/minute) passed/minute) Reaction Rate Reaction Rate (Beads (Beads joined /minute) joined /minute) 6 6 10 10 12 12 15 15 20 20 30 30 60 60 6 6 5 5 4 4 3 3 2 2 1 1

  7. Increasing the substrate concentration O As concentration of substrate increases, collisions between enzyme and substrate molecules occur more often. O More enzyme-substrate complexes form, so more product is formed. O The reaction rate increases Page 132-135

  8. The effect of substrate concentration on the rate of an enzyme controlled reaction 7 7 -- -- 6 6 -- -- Why has the rate levelled out? What is limiting any further increase in the rate of reaction ? Rate of reaction (au) 5 5 -- -- 4 4 -- -- 3 3 -- -- Here the increase in rate and substrate conc are proportional 2 2 -- -- 1 1 -- -- | | | | | | | | | | 1 2 3 4 5 1 2 3 4 5 Substrate conc (mol/dm ) Exam question. Exam question. Describe the relationship between the substrate concentration and the rate of the reaction. Look for overall trends Refer to points on the graph Do not generalise

  9. Increasing the enzyme concentration O As the enzyme concentration increases, more active sites become available. O More enzyme-substrate complexes form (and so more products are produced). The rate of reaction increases. Sketch a graph to show the relationship between enzyme Sketch a graph to show the relationship between enzyme concentration and the rate of reaction. concentration and the rate of reaction. What happens if What happens if the substrate the substrate concentration increases further? concentration increases further? Draw another line to show what the reaction would look like if Draw another line to show what the reaction would look like if there was double the substrate concentration there was double the substrate concentration 1. 1. 2. 2. 3. 3.

  10. Enzymes and temperature O Kinetic energy and collision theory O Heat, vibration, breaking bonds and denaturation Give a definition of denaturation Give a definition of denaturation Describe how heating can lead to denaturation in enzyme Describe how heating can lead to denaturation in enzyme structure structure Textbook p128-9

  11. Optimum temperatures Refer to the graph to explain the effect of temperature on enzymes. Include: Include: Kinetic energy, Collision theory , enzyme/substrate complexes, Kinetic energy, Collision theory , enzyme/substrate complexes, denaturation denaturation Textbook p128-9

  12. Precise terminology Precise terminology Formation of enzyme substrate complexes Increase in the kinetic energy of the substrate and enzyme molecules High temperatures disrupt the tertiary structure of the enzyme by breaking the tertiary bonds

  13. Enzymes and pH O What is pH? O pH and bonds (interfere with hydrogen bonds and ionic bonds-tertiary structure affected) Textbook p130-131

  14. pH and active site Importance in terms of induced-fit hypothesis Increasing H ions, alters charges around the active site Textbook p130-131

  15. O Varies depending on enzyme (e.g. where found) O At optimum, concentration of H ions in solution gives the tertiary structure the best overall shape. O Range is generally fairly narrow Optimum pH Buffer solutions at set pH values are used to carry out enzyme reactions. The rate can then be calculated and the optimum found. Denaturation will only occur in extreme changes of pH away from the optimum. The further from the optimum the faster the rate of denaturation will be. Textbook p130-131

  16. Inhibitors An enzyme inhibitor is any substance or An enzyme inhibitor is any substance or molecule that slows down the rate of an molecule that slows down the rate of an enzyme enzyme- -controlled reaction by affecting the controlled reaction by affecting the enzyme molecule in some way enzyme molecule in some way. . O Competitive Competitive Similar in shape to substrate O Non competitive Non competitive Do not compete with substrate molecules for a place in the active site O Permanent Permanent Most competitive inhibitors are not permanent (action is reversible). Many non competitive inhibitors bind permanently to enzyme molecules

  17. Enzyme inhibitors forward_arrow_colour

  18. S Normal enzyme substrate binding: E Irreversible inhibition O The inhibitor binds permanently with the enzyme and changes the shape of the active site preventing substrate binding. O e.g. heavy metal ions mercury, lead S Substrate no longer fits into active site and effect is irreversible E I I

  19. Reversible inhibition e.g. Leeches produce hirudin which inhibits the production of a blood clotting protein e.g. cyanide inhibits an enzyme involved in respiration O Non-competitive O Competitive S S I I E E E Inhibitor binds at another site (allosteric site) on the enzyme but changes the shape of the active site Substrate cannot bind, even if its concentration is increased Effect is however reversible if inhibitor is removed shape of active site is restored I If inhibitor occupies the active site and substrate can no longer bind Increasing concentration of substrate reduces effect of inhibitor

  20. With a competitive inhibitor With a non competitive inhibitor Rate Of reaction Rate Of reaction Normalrate Inhibited rate Substrate concentration Substrate concentration Decide which graph represents the effect on the rate of an enzyme controlled reaction when exposed to: a competitive inhibitor a non competitive inhibitor. Each reaction is carried out with a fixed quantity of inhibitor.

  21. End-product inhibition O The product of the reaction which is being catalysed acts an inhibitor of the enzyme involved substrate products 1. Substrate binds at active site 2. Reaction is catalysed 3. Product inhibits enzyme (usually non- competitive)

  22. True or false? True True Well Well Well Well False False False False O An enzyme is a biological catalyst O An enzyme will work faster at 60 C O An enzyme will be denatured at 100 C O Enzymes break things down O Enzymes are denatured if chilled, which is why a fridge preserves food O There will be as much enzyme left at the end of a reaction, as there was at the beginning O Blank-ase breaks down or interacts with blank True True True True

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