ENZYMES

A protein with catalytic properties due to its

power of specific activation

© 2007 Paul Billiet ODWS

Chemical reactions
 Chemical reactions need an initial input of energy =
THE ACTIVATION ENERGY
 During this part of the reaction the molecules are
said to be in a transition state.

© 2007 Paul Billiet ODWS

Reaction pathway

© 2007 Paul Billiet ODWS

Making reactions go faster
 Increasing the temperature make molecules move
faster
 Biological systems are very sensitive to temperature
changes.
 Enzymes can increase the rate of reactions without
increasing the temperature.
 They do this by lowering the activation energy.
 They create a new reaction pathway “a short cut”

© 2007 Paul Billiet ODWS

 An enzyme controlled pathway

 Enzyme controlled reactions proceed 108 to 1011 times faster

than corresponding non-enzymic reactions.
© 2007 Paul Billiet ODWS
Enzyme structure
 Enzymes are
proteins
 They have a
globular shape
 A complex 3-D
structure

Human pancreatic amylase

© Dr. Anjuman Begum

© 2007 Paul Billiet ODWS

The active site
 One part of an enzyme,
the active site, is
particularly important
 The shape and the
chemical environment
inside the active site
permits a chemical
reaction to proceed
© H.PELLETIER, M.R.SAWAYA
ProNuC Database more easily

© 2007 Paul Billiet ODWS

Cofactors
 An additional non-
protein molecule that is
needed by some
enzymes to help the
reaction
 Tightly bound cofactors
are called prosthetic
groups
 Cofactors that are bound
and released easily are
called coenzymes
 Many vitamins are
coenzymes Nitrogenase enzyme with Fe, Mo and ADP cofactors
Jmol from a RCSB PDB file © 2007 Steve Cook
H.SCHINDELIN, C.KISKER, J.L.SCHLESSMAN, J.B.HOWARD, D.C.REES
STRUCTURE OF ADP X ALF4(-)-STABILIZED NITROGENASE COMPLEX AND ITS
© 2007 Paul Billiet ODWS
IMPLICATIONS FOR SIGNAL TRANSDUCTION; NATURE 387:370 (1997)
The substrate
 The substrate of an enzyme are the reactants
that are activated by the enzyme
 Enzymes are specific to their substrates
 The specificity is determined by the active
site

© 2007 Paul Billiet ODWS

The Lock and Key Hypothesis
 Fit between the substrate and the active site of the enzyme is
exact
 Like a key fits into a lock very precisely
 The key is analogous to the enzyme and the substrate
analogous to the lock.
 Temporary structure called the enzyme-substrate complex
formed
 Products have a different shape from the substrate
 Once formed, they are released from the active site
 Leaving it free to become attached to another substrate

© 2007 Paul Billiet ODWS

The Lock and Key Hypothesis

S
E
E
E

Enzyme- Enzyme may

substrate be used again
complex P

Reaction coordinate
© 2007 Paul Billiet ODWS
The Lock and Key Hypothesis
 This explains enzyme specificity
 This explains the loss of activity when
enzymes denature

© 2007 Paul Billiet ODWS

The Induced Fit Hypothesis
 Some proteins can change their shape
(conformation)
 When a substrate combines with an enzyme, it
induces a change in the enzyme’s conformation
 The active site is then moulded into a precise
conformation
 Making the chemical environment suitable for the
reaction
 The bonds of the substrate are stretched to make the
reaction easier (lowers activation energy)
© 2007 Paul Billiet ODWS
The Induced Fit Hypothesis

Hexokinase (a) without (b) with glucose substrate

http://www.biochem.arizona.edu/classes/bioc462/462a/NOTES/ENZYMES/enzyme_mechanism.html

 This explains the enzymes that can react with a

range of substrates of similar types
© 2007 Paul Billiet ODWS
Factors affecting Enzymes
 substrate concentration
 pH
 temperature
 inhibitors

© 2007 Paul Billiet ODWS

Substrate concentration: Non-enzymic reactions

Reaction
velocity

Substrate concentration

 The increase in velocity is proportional to the

substrate concentration
© 2007 Paul Billiet ODWS
Substrate concentration: Enzymic reactions

Vmax

Reaction
velocity

Substrate concentration
 Faster reaction but it reaches a saturation point when all the
enzyme molecules are occupied.
 If you alter the concentration of the enzyme then Vmax will
change too.
© 2007 Paul Billiet ODWS
The effect of pH
Optimum pH values

Enzyme
activity Trypsin

Pepsin

1 3 5 7 9 11

© 2007 Paul Billiet ODWS

pH
The effect of pH
 Extreme pH levels will produce denaturation
 The structure of the enzyme is changed
 The active site is distorted and the substrate
molecules will no longer fit in it
 At pH values slightly different from the enzyme’s
optimum value, small changes in the charges of the
enzyme and it’s substrate molecules will occur
 This change in ionisation will affect the binding of
the substrate with the active site.
© 2007 Paul Billiet ODWS
The effect of temperature
 Q10 (the temperature coefficient) = the increase in
reaction rate with a 10°C rise in temperature.
 For chemical reactions the Q10 = 2 to 3
(the rate of the reaction doubles or triples with every
10°C rise in temperature)
 Enzyme-controlled reactions follow this rule as they
are chemical reactions
 BUT at high temperatures proteins denature
 The optimum temperature for an enzyme controlled
reaction will be a balance between the Q10 and
denaturation.
© 2007 Paul Billiet ODWS
The effect of temperature

Q10 Denaturation
Enzyme
activity

0 10 20 30 40 50
Temperature / °C

© 2007 Paul Billiet ODWS

The effect of temperature
 For most enzymes the optimum temperature is about
30°C
 Many are a lot lower,
cold water fish will die at 30°C because their
enzymes denature
 A few bacteria have enzymes that can withstand very
high temperatures up to 100°C
 Most enzymes however are fully denatured at 70°C

© 2007 Paul Billiet ODWS

Inhibitors
 Inhibitors are chemicals that reduce the rate of
enzymic reactions.
 The are usually specific and they work at low
concentrations.
 They block the enzyme but they do not usually
destroy it.
 Many drugs and poisons are inhibitors of
enzymes in the nervous system.
© 2007 Paul Billiet ODWS
The effect of enzyme inhibition
 Irreversible inhibitors: Combine with the
functional groups of the amino acids in the
active site, irreversibly.
Examples: nerve gases and pesticides,
containing organophosphorus, combine with
serine residues in the enzyme acetylcholine
esterase.

© 2007 Paul Billiet ODWS

The effect of enzyme inhibition
 Reversible inhibitors: These can be washed
out of the solution of enzyme by dialysis.
There are two categories.

© 2007 Paul Billiet ODWS

The effect of enzyme inhibition
1. Competitive: These
compete with the
substrate molecules for E+I EI
the active site.
Reversible Enzyme inhibitor
The inhibitor’s action is reaction complex
proportional to its
concentration.
Resembles the substrate’s
structure closely.

© 2007 Paul Billiet ODWS

The effect of enzyme inhibition

Succinate Fumarate + 2H++ 2e-

Succinate dehydrogenase

CH2COOH COOH CHCOOH

CH2

CH2COOH CHCOOH
COOH
Malonate

© 2007 Paul Billiet ODWS

The effect of enzyme inhibition
2. Non-competitive: These are not influenced by the
concentration of the substrate. It inhibits by binding
irreversibly to the enzyme but not at the active site.
Examples
 Cyanide combines with the Iron in the enzymes
cytochrome oxidase.
 Heavy metals, Ag or Hg, combine with –SH groups.

These can be removed by using a chelating agent such

as EDTA.
© 2007 Paul Billiet ODWS
Applications of inhibitors
 Negative feedback: end point or end product
inhibition
 Poisons snake bite, plant alkaloids and nerve
gases.
 Medicine antibiotics, sulphonamides,
sedatives and stimulants

© 2007 Paul Billiet ODWS