Enzymes

Nearly all the reactions of the body are mediated by enzymes.

Enzymes are protein catalysts that increase the rate of the
reactions without being changed in the overall process.
It is catalysts for biological reactions,
it directs all metabolic events in the body
Lower the activation energy
Increase the rate of reaction
Activity lost if denatured
May be simple proteins
May contain cofactors such as metal ions or organic (vitamins)

Enzymes
Nomenclature of enzymes
Enzyme has two names:
a. Short Recommended name
b. Systemic name
Recommended name
1. Adding suffix -ase, attached to the substrate of the reaction and
identifies a substance as an enzyme substrate e.g:
sucrase - reacts with sucrose and
lipase - reacts with lipid
2. Describes function of enzyme
Oxidase - catalyzes oxidation, hydrolase - catalyzes hydrolysis, lactate
dehydrogenase, adenylate cyclase.
3. Common names of digestion enzymes still in use which don't provide
any hint as pepsin and trypsin

Systematic name
The International Union of Biochemistry and Molecular Biology (IUBMB)
developed a system for nomenclature in which enzymes are divided into 6
classs, each with sub classes. These names are unambiguous and informative
but sometimes long and difficult to be of general use.
Class
1- Oxidoreductases
2- Transferases
groups
3- Hydrolases
water
4- Lyases
CN bonds
5- Isomerases
6- Ligases

Reactions catalyzed
Catalyze oxidation-reduction reactions
Catalyze transfer of C-, N-, or P-containing
Catalyze cleavage of bonds by addition of
Catalyze cleavage of CC, CS and certain
Catalyze racemization of isomers
Combine molecules using ATP

1. Oxidoreductases
Catalyze oxidation-reduction
reactions
Lactate formation in muscle: In
exercising skeletal muscle, NADH
production (by glycolysis (6) and
production of NAD+ by citric acid cycle),
exceeds the oxidative capacity of the
respiratory electron transfer chain ETC.
This results in an elevated NADH/NAD+
ratio, favoring reduction of pyruvate
to lactate. Therefore, during intense
exer-cise, lactate accumulates in muscle,
causing a drop in the intra-cellular pH,
potentially resulting in cramps. Much of
this lactate eventually diffuses into the
bloodstream, and can be used by the
liver to make glucose

Reduction of pyruvate to lactat

Transferases

alyze transfer of C-, N-, or P-containing groups

+THF-CH2+ H2O

THF +

Serine hydroxy-methyl transferase

CH2 group: C- containing group is removed by the enzyme

Tetrahydrofolate (THF)

3. Hydrolases:
Catalyze cleavage of bonds by
addition of water

NH2-C-NH2 + H2O

CO2 + 2NH3

O
Urea

Urease

4. Lyases
Catalyze cleavage of CC, CS and
certain CN bonds

3-C-COOH

uvate

CH3-CH +
Pyruvate
decarboxylase

Acetaldehy

5. Isomerases
Catalyze racemization of optical or
geometric isomers

Formation of Lmethylmalonyl CoA:

the D-isomer is converted
to the
L-isomer by the
Enzyme: methylmalonyl CoA
racemas

6. Ligases

combine molecules using ATP

Catalyze formation of bonds between carbon and C, O,

S, N coupled to hydrolysis of high-energy phosphates
(ATP)

Biotin

Biotin

Property of Enzyme:
Enzymes are protein catalysts that increase the
velocity of a chemical reaction, and are not
consumed during the reaction. A few Ribozymes
are RNAs that act like enzymes, and catalyze cellular
reactions involving nucleic acids, much less common than protein catalysts.
1. Active sites: an enzyme binds a substrate in a region called the active site
Active site is a special pocket or cleft in the enzyme molecule .
The active site contains amino acid side chains that form a three dimensional surface
complementary to the substrate. Formed by groups that come from different parts

of the protein chain(s) these groups brought together by the folding and
bending (secondary and tertiary structures).

2. Enzyme substrate (ES) complex: catalysts , in general, offer an

alternative pathway with lower energy through which a reaction can occur. In
enzyme-controlled reactions , this alternative pathway involve the formation of
an enzyme-substrate complex as an intermediate species in the reaction. Within
this ES complex orientation effects create more favorable reaction conditions
than if substrate were free. The result is faster formation of products.
The substrate binds the enzyme, forming an enzyme substrate (ES) complex,
it is causing conformational change in the enzyme (induced fit) that allows
catalysis. ES is converted to EP complex that subsequently dissociates to
E and P
dissociates

( E +S

ES complex)

(EP complex

E + P)

3. Catalytic efficiency: enzyme-catalyzed reactions are highly efficient,

proceeding from (1000-100,000,000) faster than uncatalyzed reactions. The
number of molecules of (S) converted to product (P) per enzyme molecule per
second is called the turnover number or Kcat.
The active site binds to the substrate and form enzyme-substrate complex that
will dissociate into the enzyme (E) and product (P).
Amino acid R groups in the active site help substrate bind.
4. Specificity: Only certain substrates can fit the active site of enzyme
catalyzing one type of chemical reaction.

5. Enzyme structure:
a. Simple enzyme: composed only of protein (amino acid chains).
b. Conjugated enzyme: have a non-protein portion in addition to a protein portion.
By itself, neither the protein part nor the non-protein portion has catalytic properties.
Only the combination of the two- the conjugated enzyme- shows biological activity and
called (Holoenzymes).
Holoenzymes: holoenzyme is the active enzyme with its nonprotein
component , whereas
Apoenzyme is the enzyme without its nonprotein moiety and inactive.
If the nonprotein moiety is a
metal ion such as: Zn2+ or Fe2+ Mg++, Mn++. it is called
cofactor.
organic molecule, (vitamin)
it is termed coenzyme.
Holoenzymes = Apoenzymes + nonprotein moiety =
Apoenzyme =

Holoenzymes

nonprotein moiety =

biologically active
biologically inactive

Enzymes

6. Location within the cell

Many enzymes are localized in
specific organelles within the cell.
Such compartmentalization
serves to isolate the reaction
substrate or product from other
competing reactions. This provides:
a favorable environment for the
reaction, & organizes the
thousands of enzymes present in
the cell into purposeful pathways.

Models of enzyme action

1. Lock and Key Model: to account for the highly specific way enzymes select a substrate and bind it to
the active site, researchers have proposed several models.
The simplest of these models is Lock and Key Model.
In the lock and key model, the active site in the enzyme has a fixed, rigid geometrical conformation.
Only substrates with a complementary geometry can be accommodated at such a site, much as a lock
accepts only certain keys.

2. Induced fit model: the lock and key model explains the action of many enzymes. It is,
however, too restrictive for the action of other enzymes. The induced fit model gives more
thorough explanation for the active site property of an enzyme because it includes the
specificity of the lock and key model coupled with the flexibility of the enzyme protein.
The forces brigs the substrate into the active site are:
- Ionic attractions
- Hydrogen bonds
- Hydrophobic interactions
For example: a positively charged amino group in a substrate could be attracted and held
at the active site by negatively charged aspartate or glutamate residue. Alternatively,
cofactors such as positively charged metal ions help bind substrate molecules

Enzymes

Enzyme specificity: enzymes exhibit different levels of selectivity, or specificity, for

substrates. This is determined by the active site. Some active sites accommodate
only one particular compound, whereas others can accommodate a family of
closely related compounds.
Types of enzyme specificities:
1. Absolute specificity: Such specificity means an enzyme will catalyze a particular
reaction for only one substrate. This is most restrictive but not common specificity.
(example Urease: will act on urea only).
2. Stereochemical specificity: here an enzyme can distinguish between stereoisomers.
(L-Amino oxidase will catalyze reactions of L-amino acid but not D-amino acids).
3. Group specificity: this specificity involves structurally similar compounds that
have the same functional groups. (Carboxypeptidase is group-specific; it cleaves
amino acids, one at a time, from the carboxyl end of the peptide chain).
4. Linkage specificity: it involves a particular type of bonds. (Phosphatase
hydrolyze phosphate-ester bonds in all types of phosphate esters).

Factors Affecting Enzyme Action

Enzyme activity is a measure of the rate at which an enzyme convert
substrate to product.
Different enzymes show different response to changes in substrate
concentration, temperature, enzyme concentration and pH
1. Temperature
Little activity at low temperature. Rate increases with increasing temperature
(the velocity increased with Tem until a peak due to the increased number of
molecules having sufficient energy to pass over the energy barrier and form
product)
Most active at optimum temperatures (usually 37 C in humans). Activity lost
with denaturation at high temperatures, the velocity decrease.

2. pH: hydrogen ion concentration: the pH of an enzyme environment can

affect its activity. The charge on acidic and basic amino acids located at the
active site depends on pH. Small change in pH (less than one unit) can result in
enzyme denaturation and subsequent loss of catalytic activity.
Optimum pH is the pH at which an enzyme has maximum activity. Each
enzyme has characteristic optimum pH, which usually falls within the
physiological pH range of 7-7.5.
Exceptions are: pepsin: working in the stomach at pH of 2.0
tripsin: working in the intestine at pH of 6.0
alkaline phosphatase
at pH of 9.0
A variation from normal pH can also affect substrates, causing either
protonation or deprotonation of groups on the substrate. The interaction
between the altered substrate and enzyme active site become less efficientor even impossible.
Optimum pH is the pH at which an enzyme has maximum activity

Each enzyme has

its optimum pH

2.0

6.0

9.0

3. substrate concentration:
Reaction Rate (velocity of a reaction): micro- moles / minute
Reaction rate is the number of substrate molecules converted to product per
unit time and is usually expressed as micro- moles of product formed per
minute. The rate of reaction (enzyme catalyzed) increases with substrate
concentration until a maximal velocity (Vmax), is reached. the plateau
reflects the saturation with a substrate of all available binding sites on the
enzyme .
When each enzyme molecule is working at full capacity, the incoming
substrate molecules must wait their turn for an empty active site. At this
point, the enzyme is said to be under saturation condition.
Turnover number: is equal to the number of substrate molecules
transformed per second by one molecule of enzyme under optimum
condition of temperature, pH, and saturation.
Maximum activity
Increasing substrate concentration increases the rate of reaction (enzyme
concentration
is constant)
Maximum activity
reached when all of enzyme molecules combines with substrate
Maximum activity reached when all of enzyme molecules combines with substrate

enzyme is said to be under saturation condition

Enz. Maximum activity

micro- moles / minute

the number of substrate molecules

converted to product per unit time

Enzyme
under saturation condition

a maximal velocity (Vmax),

Constant:
pH
Temp
Enz. Concent.

4. Enzyme concentration: the cell usually keeps the number

of enzymes low compared with the number of substrate
molecules. This is efficient; the cell avoid paying the energy
cost of synthesizing and maintaining a large work force of
enzyme molecules. Thus, in general, the concentration of
substrate in a reaction is much higher than that of the
enzyme.
If the amount of substrate present is kept constant and the
enzyme concentration is increased, the reaction rate increases
because more substrate molecule can be accommodated in
given amount of time.
the concentration of substrate in a reaction is much higher than that of the enzyme.

substrate present is kept constant

Enzyme inhibition:
The rate of enzyme-catalyzed reaction decreased by a group of substances
called inhibitors. An enzyme inhibitor is a substance that slows or stops the
normal catalytic function of an enzyme by binding to it. Three modes by
which inhibition takes place:
1. Reversible competitive inhibition:
A competitive enzyme inhibitor is a molecule that sufficiently resembles an
enzyme substrate in shape and charge distribution that it can compete
with the substrate for occupancy of the enzymes active site.
When a competitive inhibitor binds to an enzyme active site, the inhibitor
remains unchanged (no reaction occurs), but its physical presence at the
site prevents a normal substrate molecule from occupying the site. The
result is a decrease in enzyme activity. Enzyme-inhibitor (EI) complex is
a reversible process. Because it is maintained by week interactions.
When inhibitor and substrate molecules are in the vicinity they compete
for the active site of an enzyme. If the inhibitor concentration is greater
than substrate concentration, the inhibitor dominates the occupancy
process. The reverse is also true.

2-Statin drugs:
-Atorvastatin
(lipitor)
-Pravastatin
(Pravachol)

3-Hydroxy-3-methylglutaryl CoA
Hydroxy
Methylglutaryle
CoA-Reductase
Mevalonic acid

-Atorvastatin
(lipitor)

Pravastatin

Competes to inhibit
HMG-CoA
Reductase and
inhibit cholesterol
synthesis
Cholesterol

3- Sulpha Drugs: are structural analogues of para amino benzoic acid

(PABA)

Sulpha drugs: are structural analogues of para amino benzoic acid (PABA) -3
Sulfanilamide and Trimethoprim
F.A
Folic acid
Synthetas
e

F.A.Reductase

Dihydro F.A.Reduct
Amino acid synth

para amino benzoic acid

Purine synth
Pteridine + PABA + Glutamic acid

Folic Acid

Dihydro

Tetrahydro folic acid

Precursor
Inhibited by
Sulfanilamide

Inhibited by

Thymidine
Mono
Phosphate
synth

Trimethoprim

PABA: Para amino benzoic acid

F.A:
Folic acid
These drugs interfere in the synthesis of folic acid( an essential ingredient for DNA and RNA synthesis.
Used in combination. Now confined to infection in HIV-infected persons.

5- Angiotensin-converting enzyme (ACE) inhibitors: (lowers blood pressure)

Angiotensin-converting
enzyme (ACE)

Angiotensin

Angiotensin-I

inhibitors:
-Captopril
-Enalapril
-Lisinopril

Angiotensin-II

Cleaves Angiotensin-I
to form Angiotensin-II

Potent
vasoconstrictor

When the enzyme is

blocked by the drugs
the effect will be
opposite to the effect
of Angiotensin-II, so
drugs will cause
vasodilation lowering
blood pressure

2. Reversible noncompetitive inhibition:

A noncompetitive inhibitor is a molecule that decrease enzyme activity by
binding to a site on an enzyme other than the active site(allosteric site).
In the process of binding to the enzyme, noncompetitive inhibitors
usually alter the overall tertiary structure of the enzyme, including the
structure at the active site. Changing the shape of the active site lower
the affinity of the enzyme for its substrate and/or the enzymes overall
ability to function. Lowering the concentration of a noncompetitive
inhibitor sufficiently does free many enzymes, which return to normal
activity.
Examples: the heavy metal ions Pb ++, Ag+, Hg++ . The
binding sites for these ions are:
sulfhydryl (-SH) groups of cystein of the enzyme
LOCATED AWAY FROM THE ACTIVE SITE. Metal
disulfide linkage are formed which disrupts the
secondary and tertiary structure.

2. Reversible noncompetitive inhibition

Changing the shape of the active site

lower the affinity of the enzyme
for its substrate and/or the enzymes
overall ability to function

LOCATED AWAY FROM THE ACTIVE SI

sulfhydryl (-SH) groups

The binding sites for these

ions are:
sulfhydryl (-SH) groups.
Metal disulfide linkage are
formed which disrupts the
secondary and tertiary
structure.

3. Irreversible inhibition:
An irreversible enzyme inhibitor: is a molecule that inactivates enzymes by
forming a strong covalent bond to an amino acid-side chain group at the
enzyme active site. Such inhibitors do not have structures similar to that
of enzymes normal substrate. The inhibitor-active site bond is
sufficiently strong that addition of excess substrate does not reverse the
inhibition process. Thus the enzyme is permanently deactivated.
examples: the action of chemical warfare (nerve gases) and
organophosphate insecticides are based on irreversible inhibition

3. Irreversible inhibition

Organophosphate compounds
Irreversible enzyme inhibitors

The phosphate group in a nerve gas irreversibly

attach to OH of serine to form phosphoester ,
this blocks the enzyme active site
phosphate
Gas DIPFP is irreversible enzyme inhibitors
group

irreversibly attach to OH of seri

DIPFP

The R
of
serine
of
the
enzy
me

Covalent bond blocks the active site of the enzyme acetylcholinesterase

from substrate use. (acetylcholine)

Regulation of enzyme activity

A. Allosteric enzymes:
Many of the molecules responsible for regulating cellular processes are special group of enzymes
called allosteric enzymes.
Allosteric enzyme: is a molecule with 2 or more chains (quaternary structure) and 2 kinds of binding
sites (substrate B.S and regulator B.S).
Substances that bind at regulatory sites of allosteric enzymes are called:
regulators. (effectors or modifiers).
The binding of a positive regulator increases enzyme activity; the shape of the active site is changed
such that it can more readily accept substrate.
The binding of a negative regulator (noncompetetive inhibitor) decreases enzyme activity; the shape
of the active site is changed such that it can less readily accept substrate.
1. Homotropic effectors: when the substrate itself serves as an effector, the effect is said to be
homotropic. Most often, an allosteric substrate function as a positive effector. In such a case the
presence of a substrate molecule at one site on the enzyme enhances the catalytic properties of the
other substrate-binding sites- that is, their binding sites exhibit cooperativity.

bind here

Allosteric enzymes has

2 binding sites (B.S)

substrate B.S and regulator B.S

Regulators
(effectors or modifiers)

Positive Regulators
The binding of a positive regulator increases enzyme activity;
the shape of the active site is changed such that it can more readily accept substrate.
Negative Regulators
The binding of a negative regulator decreases enzyme activity;
the shape of the active site is changed such that it can less readily accept substrate.

2. Heterotropic effectors: the effector may be different from the substrate, in which case
the effector is said to be heterotropic. For example, consider the feed back inhibition
control shown bellow:
(D) is a negative regulator
of the 1st enzyme
Enzyme 1
A

Enzyme 2
B

Enzyme 3
C

The above biochemical process in a cell occurs in several steps, each step catalyzed by a
different enzyme. The product of each step is the substrate for the next enzyme.
If the final product (D) is a negative regulator of the 1st enzyme (enzyme 1) .
-At low concentration of D, the reaction sequence proceeds rapidly.
-At higher concentration of D, the activity of enzyme 1 becomes inhibited (by feed back),
and eventually the activity stops. At stopping point, there is sufficient D present in the
cell to meet its needs.
Later, when the concentration of D decreases through use in other reactions, the activity
of enzyme 1 increases and more D is produced.
Heterotropic effectors are commonly encountered, for example, the glycolytic enzyme
phosphofructokinase-1 is allosterically inhibited by citrate, which is not a substrate for
the enzyme.

Glucos
e
glucose 6-phosphate

fructose 6-phosphate
Phosphofructokin
ase-1

Citra
te

fructose 1,6-bisphosphate

Glycolytic pathway Pyruvate

AcetylCoA

Citrate

B. Covalent modification - Protein phosphorylation

-Many enzymes may be

regulated by covalent
modification; by addition or
removal of phosphate group
from specific -OH of serine,
threonine or tyrosine residues of
enzyme.

Inactive or

1- Phosphorylation and dephosphorylation:

Phosphrylation reactions are catalyzed by a family of enzymes called protein kinases that use
ATP as a phosphate donor. Phosphate groups are cleaved from phosphorylated enzymes by the
action of phosphoprotein phosphatases.

2- Response of enzyme to Phosphorylation:

-Phosphorylation of Glycogen Phosphorylase Enzyme increase its activity
-Phosphorylation of Glycogen synthase Enzyme decrease its activity
Phosphorylation of

glycogen phosphorylas Enzyme

Increase its activity

Degredation
To glucose

Phosphorylation of
glycogen synthase Enzyme

decrease its activity

Glycogen

Glucose

Glycogen
Synthesis

C. Induction and repression of enzyme

synthesis

Cells can regulate the amount of enzyme present, usually by altering the
rate of enzyme synthesis. The (increase) induction or (decrease) repression
of enzyme synthesis leads to an alteration in the total number of active sites.
Enzymes subject to regulation of synthesis are often those that are
needed at only one stage of development or
under selected physiologic conditions.
Example: elevated level of insulin as a result of high blood glucose level
cause an increase in the synthesis of key enzymes involved in glucose
e.g: glucokinase adds phosphate to glucose
metabolism.
In contrast, enzymes that are in constant use are usually not regulated by
altering the rate of enzyme synthesis.
Alteration in enzyme levels as a result of induction or repression are slow
(hours to days)
Allosterically or covalently regulated changes in enzyme activity is rapid
(occur in seconds to minutes)

D. Regulation of enzyme activity: zymogens

Because they would otherwise destroy the tissues that produce them,
some enzymes are generated in an inactive form and then converted to an
active form when it is needed. Most digestive and blood-clotting enzymes
are of this type. These inactive enzymes are zymogens or proenzymes.
(inactive precursor of an enzyme).
Activation of a zymogen requires either adding to or removing some
parts of it. This modification will change the three-dimensional structure
(secondary and tertiary) of the zymogen, which affect the active site
conformation. Example: the zymogen pepsinogen is converted to the
active enzyme pepsin in the stomach, then function as digestive enzyme.
Pepsin activation involves removal of a polypeptide fragment from its
structure.
inactive enzymes are zymogens or proenzymes. (inactive precursor of an
enzyme).

digestive and blood-clotting enzymes

Inactive: zymogen

Active: zymogen

zymogen pepsinogen

Activated pepsin

removal of a polypeptide fragment

Medical uses of enzymes:

Enzymes can be used to diagnose certain diseases. Although blood plasma
contains many enzymes, some enzymes are not normally found in the blood
but are produced only inside cells of certain organs and tissues. The
appearance of these enzymes in the blood often indicates that there is tissue
damage in an organ and that cellular contents are spilling out (leaking)
into the blood stream. Assays of abnormal enzyme activity in blood serum
can be used to diagnose many disease states, some of which are listed in the
next table, also enzymes that are used to diagnose heart attacks
(myocardial infarctions) (M.I) are shown the next slide.
CK2 (MB), following an acute M.I, this enzyme appears approximately 4-8
hours following onset of chest pain, reaches a peak of activity at 24 hours,
the return to baseline after 48-72 hours.
Troponin T and Troponin I appears in plasma within 4-6 hours after an
M.I, peaks in 8-28 hours, and remains elevated for 3-10 days.

Cardiac tissue damage

Enzyme leakage

Creatine phosphokinase
CP K 1,2,3
- Troponin T and
- Troponin I

OR

Creatine Kinase
CK2 (MB)

Brain: CKI
= BB
Cardiac: CK2 =MB
Skeletal: CK3 = MM

Tissue plasminogen activator (TPA) , which activates the

enzyme plasminogen is used in the treatment of heart attack .
This enzyme when activated dissolves blood clots in the heart
and often provides immediate relief.
Normal

Cell damage

less than 1

LDH 1/LDH 2 Ratio

Greater than 1

<1
>1

<1

Vitamins
Many enzymes contain vitamins as part of their structure. Conjugated
enzymes have a protein part (apoenzyme) and a nonprotein part (cofactor).
Vitamins, in many cases, are cofactors in conjugated enzyms.
Vitamins: are organic compounds that must be obtained from dietary
sources and that are essential in trace amounts for the proper functioning
of the human body. Vitamins must be obtained from the diet, because the
human body cannot synthesize them in the required amounts. Needed in
microgram or milligram quantities per day compared with 50-200 grams
per day for the major classes of foods (carbohydrates, lipids and protein) .
The recommended daily allowance (RDA) of vitamin B12 is 2.0
micrograms /day for an adult. Just 1.0 gram of it could supply the daily
needs of 500,000 people. A well-balanced diet usually meets all the bodys
vitamin requirements.

Conclusions about Michaelis - Menten kinetics

Characteristics of Km :
The Km constant is a characteristics of an enzyme and
particular substrate
Km reflects the affinity of the enzyme for a particular
substrate
Km numerically equal to the concentration substrate at
which the reaction velocity is equal to Vmax.
Km dose not vary with concentration of the enzyme
low Km high affinity; low [S] is needed to halfsaturate the enzyme
large Km low affinity;
high [S] is needed to half-saturate the enzyme

A. Reaction model
Leonor Michaelis and Maude Menten
proposed a simple model that accounts for
most of the features of enzyme-catalyzed
reactions. In this model, the enzyme
reversibly combines with its substrate to
form an ES complex that subsequently
yields product, regenerating the free
enzyme. The model, involving one substrate
k1
k2
molecule, is represented below, where
k-1

E + S

ES

E + P

S is the substrate
E is the enzyme
ES is the enzymesubstrate complex
P is the product
k1, k-1, and k2 are rate constants

The Michaelis-Menten equation

describes how reaction velocity varies
with substrate concentration:
Where:

Vo = initial reaction velocity

Vmax = maximal velocity
Km = Michaelis constant = (k-1
+ k2)/k1
[S] = substrate concentration
The following assumptions are
made in deriving the MichaelisMenten rate equation:

1. Relative concentrations of E and S:

The concentration of substrate ([S]) is
much greater than the concentration of
enzyme ([E]), so that the percentage of
total substrate bound by the enzyme at
any one time is small.
2. Steady-state assumption: [ES] does
not change with time (the steady-state
assumption), that is, the rate of formation
of ES is equal to that of the breakdown of
ES (to E + S and to E + P). In general, an
intermediate in a series of reactions is said
to be in steadystate when its rate of
synthesis is equal to its rate of
degradation.
3. Initial velocity: Initial reaction
velocities (Vo) are used in the analysis of
enzyme reactions. This means that the
rate of the reaction is measured as soon as
enzyme and substrate are mixed. At that
time, the concentration of product P is
very small and, therefore, the rate of the

IMPORTANT CONCLUSIONS ABOUT MICHAELISMENTEN KINETICS

1. Characteristics of Km: Kmthe Michaelis
constantis characteristic of an enzyme and its
particular substrate, and reflects the affinity of the
enzyme for that substrate. Km is numerically equal
to the substrate concentration at which the reaction
velocity is equal to 12Vmax. Km does not vary with
the concentration of enzyme.
a. Small Km: A numerically small (low) Km reflects
a high affinity of the enzyme for substrate, because
a low concentration of sub-strate is needed to halfsaturate the enzymethat is, to reach a
velocity that is 12Vmax (Figure 5.9).
b. Large Km: A numerically large (high) Km reflects
a low affinity
of enzyme for substrate because a high
concentration of sub-strate is needed to half-saturate
the enzyme
2. Relationship of velocity to enzyme
concentration: The rate of the reaction is directly
proportional to the enzyme concentration at all
substrate concentrations. For example, if the enzyme
concentration is halved, the initial rate of the
reaction (Vo), as well as that of Vmax, are reduced
to half that of the original.
3. Order of reaction: When [S] is much less than

A. Competitive inhibition
This type of inhibition occurs when the inhibitor binds reversibly to the same site
that the substrate would normally occupy and, there-fore, competes with the
substrate for that site.
1. Effect on Vmax: The effect of a competitive inhibitor is reversed by increasing
[S]. At a sufficiently high substrate concentration, the reaction velocity
reaches the Vmax observed in the absence of inhibitor.
2. Effect on Km: A competitive inhibitor increases the Km for a given substrate.
This means that, in the presence of a competitive inhibitor, more substrate is
needed to achieve 12Vmax.

B. Noncompetitive inhibition This type of inhibition is recognized by its characteristic

effect on Vmax. Noncompetitive inhibition occurs when the inhibitor and substrate bind
at different sites on the enzyme. The noncompetitive inhibitor can bind either free
enzyme or the ES complex, thereby pre-venting the reaction from occurring.
1. Effect on Vmax: Noncompetitive inhibition cannot be overcome by increasing the
concentration of substrate. Thus, noncompetitive inhibitors decrease the Vmax of
the reaction.
2. Effect on Km: Noncompetitive inhibitors do not interfere with the binding of
substrate to enzyme. Thus, the enzyme shows the same Km in the presence or
absence of the noncompetitive inhibitor.

The End