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Enzymes
Biological Catalysts
Nomenclature and Classification
Enzyme-Substrate Interaction
Effects of pH and Temperature
Regulation of Enzyme Activity
Cofactors and Coenzymes
Vitamins and Coenzymes
Ch106; Chpt 21 Enzymes
20 - 1
Enzymes = Biological catalysts
Large proteins
Permit reactions
to ‘go’ at
body conditions
pH 7.4, 37oC
Process millions
of molecules
every second
Very specific
react with only
1 or a few types
of molecules
(substrates).
Ch106; Chpt 21 Enzymes
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Effect of enzymes on Eact
Energy
Ea
For all reactions
you must get over
the activation
energy hurdle.
Reactants
H2O2
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H
Products
H2O + O2
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Effect of enzymes on Eact
Enzyme catalyzed
reaction
Enzymes change
how reactions
proceed.
Energy
Ea
Reactants
H2O2
Ch106; Chpt 21 Enzymes
H
Reducing
activation energy.
Makes faster.
Products
H2O + O2
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Enzyme nomenclature
Name is based on:
what with or
how
it reacts
+
-ase
ending
Examples
To react with lactose.
lactase
To remove carboxyl from pyruvate.
pyruvate decarboxylase
Ch106; Chpt 21 Enzymes
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Classification of enzymes
Based on type of reaction
Oxireductase catalyze a redox reaction
Transferase
transfer a functional group
Hydrolase
catalyze hydrolysis rxns
Lyase
Add or remove to C=C bonds
Isomerases
rearrange to form isomers
Ligase
join two molecules
Ch106; Chpt 21 Enzymes
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The Active Site
Enzymes are typically HUGE proteins, yet only a
small part are actually involved in reaction.
The active site has two
basic components.
catalytic site
binding site
Model of
trios-p-isomerase
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The Active Site
Catalytic site
Binding
Site
Where reaction
occurs
holds
substrate
in
place
Substrate
Enzyme
Ch106; Chpt 21 Enzymes
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SPECIFICITY
• Enzymes are very specific. Each enzyme will
catalyze only one type of reactions and often
will only work with a specific substrate.
Ex. NH2-C-NH2 + H20
urease
2NH3 + CO2
O
• Urease has no effect on other compounds.
• Such absolute specificity is rather rare among
enzymes.
Ch106; Chpt 21 Enzymes
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Enzyme classes
Absolutely specific
Only reacts with a single substrate.
Group specific
Works with similar molecules with the same
functional group.
Linkage specific
Catalyzes a specific combination of bonds.
Stereochemically specific
Only will work with the proper D- or L- form.
Ch106; Chpt 21 Enzymes
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• ISOENZYMES: Different enzymes that perform
the same type of function in different
organisms or tissues.
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Enzyme-substrate complex
Step 1: (All of these steps are in equilibrium)
Enzyme and substrate combine to form complex
E
+
S
ES
Enzyme
Substrate
Complex
+
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Enzyme-product complex
Step 2:
An enzyme-product complex is formed.
ES
ES
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EP
transition
state
EP
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Product
The enzyme and product separate
EP
E + P
The product
is made
EP
Ch106; Chpt 21 Enzymes
Enzyme is
ready
for
another
substrate.
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Lock and Key Theory
Enzyme is “lock” and Substrate is the “key”.
Substrate structure
must fit into enzyme’s structure.
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Induced Fit Theory
Active site may not fit substrate.
Site must change in order to form the complex.
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Effect of Temp on Enzymatic Rxns
Exceeding normal pH and temperature ranges
always reduces enzyme reaction rates.
Reaction Rate
Optimum Temp
usually
37oC.
Temperature
Ch106; Chpt 21 Enzymes
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Effect of pH on Enzymatic Rxns
Reaction Rate
Most enzymes
work best near pH 7.4
not all though.
pH
Ch106; Chpt 21 Enzymes
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Examples of optimum pH
Enzyme
Source
pepsin
sucrase
catalase
arginase
alkaline
phosphatase
gastric mucosa
intestine
liver
beef liver
bone
Ch106; Chpt 21 Enzymes
Optimum
pH
1.5
6.2
7.3
9.0
9.5
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Effect of substrate concentration
Rate of reaction
(velocity)
For non-enzyme catalyzed reactions
Rate increases if
concentration of
the substrate increases
Substrate concentration
Ch106; Chpt 21 Enzymes
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Effect of substrate concentration
Rate of reaction
(velocity)
For Enzyme catalyzed reactions
Rates increase but only to a certain point
Saturation point
Vmax w/ more enzyme
Vmax w/ some enzyme
At Vmax
the enzyme is working as
fast as it can.
Rate is limited by
the concentration of both
the substrate and enzyme.
Substrate concentration
Ch106; Chpt 21 Enzymes
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Effect of Enzyme concentration
Enzyme
Activity
Enzyme Concentration
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Turnover Number
Turnover Number:
• The rate at which an enzyme transforms the
substrate
• Is measured at optimum pH and temperature.
Example:
Carbonic Anhydrase
H2CO3
H2O + CO2
36,000,000 molecules
minute
Ch106; Chpt 21 Enzymes
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ENZYME INHIBITION
• Inhibitors = interfere with ability of enzyme to
react properly with its substrate.
For example:
– Medicinal drugs
• inhibit by inactivating an enzyme
essential to bacterial growth.
Viruses more difficult to inhibit because they
use enzyme system of the host cell.
(An inhibitor of a virus also
destroys host cells)
Ch106; Chpt 21 Enzymes
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ENZYME INHIBITION
• Two Types of Inhibitors:
– Competitive
– Noncompetitive
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COMPETITIVE INHIBITOR
Competes with
substrate for
the active site.
Enzyme
mistakes
inhibitor for
substrate
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Reversible Competitive inhibition
Enzyme - substrate reactions in equilibrium.
Inhibitor
Substrate
EI
EI
shifts
ES
I + E + S
Inc I
Inc S
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ES
EP  E + P
Shifts
20 - 30
Competitive Inhibitors
Sulfa Drugs
• Illnesses caused by invading microorganisms
like bacterium can be combated using a
competitive inhibitor called an antimetabolite.
• Folic Acid is a coenzyme in many biosynthetic
processes like synthesis of amino acids and
nucleotides.
Ch106; Chpt 21 Enzymes
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O
Sulfa Drugs
C
Folic Acid : obtained
• from the diet or
OH
H N
p-aminobenzoic acid
H
• from microorganisms in the intestinal tract.
Microorganisms make folic acid from PABA.
H2N
N
N
N
N
OH
O
O
NH
C NH CH C OH
CH2
CH2
C OH
O
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Penicillin: War of Enzyme
against Enzyme.
• Produced by mold, it prevents growth of
bacteria by successfully competing for active
sites on an enzyme that bacteria need for cell
wall production.
1. Bacteria need the enzyme transpeptidase to
make their cell walls rigid and cross-linked.
2. Penicillin takes control of transpeptidase.
3. Bacteria cell walls are not cross-linked and
the contents of the bacteria cells cannot be
held in.
4. Cytoplasm spills out, and the bacteria die.
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By changing the R group, science has found a
way to prevent this from happening.
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Non competitive Inhibition
• This type of inhibitor is believed to alter the
shape of the enzyme and greatly reduce its
affinity for the substrate.
1. It does not compete with the substrate for
the active site.
2. It does not need to resemble the structure
of the substrate.
3. Its’ effect cannot be reversed by increasing
the substrate concentration.
Ch106; Chpt 21 Enzymes
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Non competitive Inhibition
A noncompetitive inhibitor
can bind to an enzyme in
many ways.
If it binds somewhere on
the surface of the enzyme,
it causes a change in the
tertiary structure.
The substrate is inhibited
because it can’t get into the
enzyme.
Ch106; Chpt 21 Enzymes
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Regulation of enzyme activity
Enzymes are often
regulated by the cell.
(Unlike other catalysts)
Cells use several
methods to control
when & how well
enzymes work.
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PROENZYMES (ZYMOGENS)
Enzymes manufactured in inactive form.
Activated when small part
of polypeptide chain removed.
Hormones,
Digestive Enz,
Blood Clotting Enz
Ch106; Chpt 21 Enzymes
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PROENZYMES (ZYMOGENS)
Enzymes manufactured in inactive form.
In pancreas (inactive)
Proinsulin
S
S
S
S
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In blood (active)
Insulin
S
S
S
S
S
S
S
S
20 - 43
PROENZYMES (ZYMOGENS)
(inactive)
In pancreas
Trypsinogen
(active)
In Intestines
enteropeptidase
Chymotrypsinogen
Trypsin
Trypsin
Chymotrypsin
procarboxypeptidase Trypsin Carboxypeptidase
Digestive
Enzymes
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Proteases
Cleave peptides
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PROENZYMES (ZYMOGENS)
(inactive)
In pancreas
Trypsinogen
(active)
In Intestines
enteropeptidase
Chymotrypsinogen
Trypsin
Trypsin
Chymotrypsin
procarboxypeptidase Trypsin Carboxypeptidase
Activation in pancreas rather than intestines 
•pancreas proteins get digested
•pancreatitis (inflammation of pancreas).
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PROENZYMES (ZYMOGENS)
(inactive)
In Gastric mucosa
Pepsinogen
Digestive
Enzyme
(active)
In Stomach
H+
Pepsin
HCl
Produced as
Food enters stomach
As pH  acid
•Proenzyme gets cleaved
•Pepsin gets activated
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Allosteric Enzymes
• Application of non competitive inhibition
• Regulates away from active site
Inactive
Enzyme
Active
Enzyme
Substrate
Now fits
Positive
Active Site
Regulator
Changed
Positive allosterism - activates the enzyme.
Negative allosterism - deactivates the enzyme.
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Inactive
Enzyme
Negative
Regulator
Feedback Control
E1
End Product
Stops E1
B
E1
Active
Allosteric
Enzyme
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E1
A
C
E2
D
E3
20 - 50
Cofactors
Apoenzyme
•protein portion
•Inactive
Co2+
Cofactor
Non protein Group
need to ‘activate’
apoenzyme
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Co2+
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Mineral Cofactors
Metal Ion
Enzyme involved
Function
Cu2+
Cytochrome oxidase
redox
Fe2+/Fe3+
Catalase
Cytochrome oxidase
redox
Zn2+
Alcohol dehydrogenase
Used with NAD+
Mg2+
Glucose-9-phosphatase Hydrolyzes
phosphate esters
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Coenzymes
Organic molecule that temporarily binds to
apoenzyme in order for it to work
+
apoenzyme
Protein
Ch106; Chpt 21 Enzymes
coenzyme
Non-Protein
holoenzyme
Total
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Vitamins are often
converted to coenzymes
Vitamin
Coenzyme made
Function
B1
thiamine pyrophosphate decarboxylation
B2
flavin mononucleotide
carries hydrogen
folic acid
tetrahydrofolic acid
amino acid
metabolism
biotin
biocytin
CO2 fixation
pantothenic Coenzyme A
acid
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acyl group carrier
20 - 59
Enzymes in Medical Diagnosis
and Treatment
• Most enzymes are confined within the cells of
the body.
• However, small amounts can also be found in
body fluids (blood, urine, cerebrospinal fluid)
• The level of enzyme activity outside the cells
can be easily monitored.
• Abnormal activity (high or low) of particular
enzymes in various body fluids signals either
the onset of certain diseases or their
progression.
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EXAMPLES
• Dead heart muscle cells spill their enzyme
contents into the serum.
• The level of glutamate oxaloacetate
transaminase (GOT) in the serum rises rapidly
after a heart attack.
• The levels of GOT as well as lactate
dehydrogenase and creatine phosphokinase
are closely monitored in order to diagnose the
severity of a myocardial infarction.
Ch106; Chpt 21 Enzymes
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Specific enzyme examples
Let’s look at role of some specific enzymes.
Two good examples are:
Chymotrypsin
- A proteolytic enzyme (protein-cleaving).
- Used in digestion of dietary protein in the
small intestines.
Acetylcholinesterase
- Used for hydrolysis of acetylcholine.
- Needed for operation of nerves.
Ch106; Chpt 21 Enzymes
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Chymotrypsin
This enzyme is a proteolytic
enzyme. It cleaves peptide
bonds.
H O
H
| ||
|
-C-HN - C - C - N - C |
| |
R
H R1
Peptide bond
Ch106; Chpt 21 Enzymes
This enzyme only works
on amino acids containing
an aromatic ring.
phenylalanine, tyrosine
and tryptophan.
20 - 64
Acetylcholinesterase and
nerve transmission
This enzyme is needed to transmit a nerve
signal at a neuromuscular junction.
Arrival of a nerve signal causes Ca2+ levels to
increase.
This causes acetylcholine containing vesicles
to move to end of the nerve cell where it is
released.
Acetylcholine then diffuses across synapse to
pass the signal to the muscle.
Acetylcholinesterase then destroys the
acetylcholine to stop the signal.
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Acetylcholinesterase and
nerve transmission
synaptic
cleft
Presence of
acetylcholine at receptor
causes a flow of sodium
and potassium ions.
This causes a muscle
contraction.
acetylcholine
receptor protein
acetylcholinesterase
- destroys excess
acetylcholine
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Acetylcholinesterase
Stick model of
acetylcholinesterase.
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Acetylcholinesterase and
nerve transmission
Without the enzyme, muscles would continue to
contract causing spasms.
Acetylcholinesterase inhibitors are used as
drugs and poisons.
Organo fluorophosphates
- bind to the enzyme. Death can occur.
Succinylcholine
Acts like acetylcholine and binds to sites on the
muscle. Used as a muscle relaxant.
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Another example
Blood Clotting - formation of fibrin.
Process requires a series of enzymatic steps.
Many of the enzymes are made in an inactive
form. This prevents blood from clotting on its
own.
Two pathways can be used to start the process.
Extrinsic
- Activated by tissue damage,
outside the blood vessel.
Intrinsic
Ch106; Chpt 21 Enzymes
- Activated by damage within a
blood vessel.
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Fibrin
Ribbon model
of fibrin.
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Drug interactions
Drugs can be administered to alter the clotting
mechanism.
Example: Heparin - an anticoagulant.
Acts by accelerating the action of the existing
inhibitor of thrombin - antithrombin III.
Antithrombin III inhibits activation of the
clotting factors that have a reactive serine
residue at their enzymatically active centers.
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thrombin
Ch106; Chpt 21 Enzymes
antithrombin
20 - 73
Heparin interaction
thrombin
serine
antithrombin
inhibited thrombin
lysine sites
heparin
Addition of heparin makes it easier for trombin to interact
with antithrombin - positive allosteric effect.
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Defective enzymes
and disease
A number of hereditary diseases result from the
absence of an enzyme or a defective one.
Disease
Defective enzyme
Albinism
tyrosinase
Glactosemia
glactose 1-phosphate
uridyltransferase
Phenylketonuria
(PKU)
phenylalanine hydroxylase
Tay-Sachs disease
hexosaminidase A
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Phenylketonuria (PKU)
• Genetic mutation that results in a defect of the
enzyme phenylalanine hydroxylase. (carried by
2% of population)
Affects about 1 baby per 13,000.
Feds may require screening at birth.
Can result in retarded physical and mental
development if untreated.
Treatment - restrict
phenylalanine until age 10
(until brain is developed).
Ch106; Chpt 21 Enzymes
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Phenylketonuria (PKU)
PKU is one of a family of enzymatic/genetic
disorders related to phenylalanine metabolism.
phenylalanine
-CH2-CH-COOH
|
NH2
tyrosine
PKU
HO-
blocked
-CH2-CH-COOH
|
NH2
albinism
melanin
alkaptonuria
O
CH-COO||
||
+
blocked
CH
C-SCoA
3
HC-COOacetyl CoA
fumarate
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-CH2-COOH
Homogentisic acid
20 - 77
ALKAPTONURIA AND
OCHRONOSIS
• Alkaptonuria is a rare disease in which the
body does not have enough of an enzyme
called homogentisic acid oxidase (HGAO)
• homogentisic acid (HGA) is not used and
builds up in the body
• Some is eliminated in the urine, and the rest is
deposited in body tissues where it is toxic.
• The result is ochronosis, a blue-black
discoloration of connective tissue including
bone, cartilage, and skin caused by deposits of
ochre-colored pigment.
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