Download Lecture 2 * The Kinetics of Enzyme Catalyzed

Lecture 2 –
The Kinetics of Enzyme Catalyzed
Reaction
Dr. AKM Shafiqul Islam
University Malaysia Perlis
01.01.10
Classification of Enzyme
• Enzymes fall into 6 classes based on function
1. Oxidoreductases: which are involved in oxidation,
reduction, and electron or proton transfer reactions
2. Transferases : catalysing reactions in which groups are
transferred
3. Hydrolases : which cleave various covalent bonds by
hydrolysis
4. Lyases : catalyse reactions forming or breaking double
bonds
5. Isomerases : catalyse isomerisation reactions
6. Ligases : join substituents together covalently.
Enzyme Reaction
• For design and analysis of a reacting system,
we must have a mathematical formula which
gives the reaction rate
in terms of
– composition,
– temperature,
– and pressure of the reaction mixture.
Enzyme Kinetics
• Enzymes are protein catalysts that, like all
catalysts, speed up the rate of a chemical
reaction without being used up in the process.
Enzyme Kinetics
• Synthetic catalysts and enzymes use the
common technique for modeling reaction
kinetics.
• The rate expressions eventually obtained for
both types of catalysts are very similar and
sometimes of identical forms.
• This is because, in both cases, the reacting
molecules form some sort of complex with the
catalyst.
Enzyme Kinetics
• Most synthetic catalysts are not specific; i.e.
they will catalyze similar reactions involving
many different kinds of reactants.
• While some enzymes are not very specific,
many will catalyze only one reaction involving
only certain substrates.
• Explain why enzyme catalysis are nonspecific
but enzyme catalysis are specific?
• Define
– Cofactors
– Apoenzyme
– Holoenzyme
– coenzyme
Enzyme Kinetic
• Enzyme is a catalyst which increases the rate
of a chemical reaction without undergoing a
permanent chemical change.
• While a catalyst influences the rate of a
chemical reaction, it does not affect reaction
equilibrium.
• Equilibrium concentrations can be calculated
using only the thermodynamic properties of
the substrates and products.
Enzyme
Enzyme Kinetics
• Both synthetic and biological catalysts can
gradually lose activity as they participate in
chemical reactions.
• However, enzymes are in general far more fragile.
Enzymes contorted shapes in space often endow
enzymes with unusual specificity and activity
It is relatively easy to disturb the native
conformation and destroy the enzyme's catalytic
properties.
Enzyme Kinetics
• It is often asserted that enzymes are more active, i.e.,
allow reactions to go faster, than nonbiological
catalysts.
• At the ambient temperatures where enzymes are most
active they are able to catalyze reactions faster than
the majority of artificial catalysts.
• When the reaction temperature is increased, solid
(synthetic) catalysts may become as active as enzymes.
• The enzyme activity does not increase continuously as
the temperature is raised. Instead, the enzyme usually
loses activity at quite a low temperature, often only
slightly above that at which it is typically found.
Enzyme Kinetics
Enzyme reaction rates are determined
by several factors.
• Concentration of substrate molecules
–
The more of them available, the quicker the enzyme molecules
collide and bind with them).
– The concentration of substrate is designated [S] and is expressed in
unit of molarity.
• Temperature.
– As the temperature rises, molecular motion - and hence collisions
between enzyme and substrate - speed up.
– But as enzymes are proteins, there is an upper limit beyond which the
enzyme becomes denatured and ineffective.
Enzymes cont.
• the presence of inhibitors.
– competitive inhibitors are molecules that bind to the
same site as the substrate - preventing the substrate
from binding as they do so - but are not changed by
the enzyme.
– noncompetitive inhibitors are molecules that bind to
some other site on the enzyme reducing its catalytic
power.
• pH. The conformation of a protein is influenced by pH and
as enzyme activity is crucially dependent on its
conformation, its activity is likewise affected.
How we determine how fast an
enzyme works
• We set up a series of tubes containing graded
concentrations of substrate, [S] . At time zero, we
add a fixed amount of the enzyme preparation.
• Over the next few minutes, we measure the
concentration of product formed. If the product
absorbs light, we can easily do this in a
spectrophotometer.
• Early in the run, when the amount of substrate is
in substantial excess to the amount of enzyme,
the rate we observe is the initial velocity of Vi.
THE ENZYME-SUBSTRATE COMPLEX
AND ENZYME ACTION
• There is no single theory which accounts for
the unusual specificity and activity of enzyme
catalysis.
• However, there are a number of plausible
ideas supported by experimental evidence for
a few specific enzymes.
• Probably, all or some collection of these
phenomena acting together combine to give
enzymes their special properties.
THE ENZYME-SUBSTRATE COMPLEX
AND ENZYME ACTION
• The x-ray crystallography, spectroscopy, and
electron-spin resonance showed the
existence of a substrate-enzyme complex.
• The substrate binds to a specific region of the
enzyme called the active site, where reaction
occurs and products are released.
• Binding to create the complex is sometimes
due to the type of weak attractive forces.
THE ENZYME-SUBSTRATE COMPLEX
AND ENZYME ACTION
• The complex is formed when the substrate key
joins with the enzyme lock.
• The hydrogen bonds formed between the
substrate and groups widely separated in the
amino acid chain of the enzyme.
THE ENZYME-SUBSTRATE COMPLEX
AND ENZYME ACTION
• The protein molecule is folded in such a way that a
group of reactive amino acid side chains in the enzyme
presents a very specific site to the substrate.
• The reactive groups encountered in enzymes include
the R group of Asp, Cys, Glu, His, Lys, Met, Ser, Thr, and
the end amino and carboxyl functions.
• Since the number of such groups near the substrate is
typically 20, only a small fraction of the enzyme is
believed to participate directly in the enzyme's active
site.
• Large enzymes may have more than one
active site.
• Many of the remaining amino acids determine
the folding along a chain of amino acids
(secondary structure) and the placement of
one part of a folded chain next to another
(tertiary structure), which help create the
active site itself
THE ENZYME-SUBSTRATE COMPLEX
AND ENZYME ACTION
• Enzymes can hold substrates so that their
reactive regions are close to each other and to
the enzyme's catalytic groups.
• This feature, which quite logically can
accelerate a chemical reaction, is known as
the proximity effect.
• Reaction will occur only when the molecules come
together at the proper orientation so that the reactive
atoms or groups are in close juxtaposition.
• Enzymes are believed to bind substrates in especially
favorable positions, thereby contributing an
orientation effect, which accelerates the rate of
reaction.
• Also called orbital steering, this phenomenon has
qualitative merit as a contributing factor to enzyme
catalysis. The quantitative magnitude of its effect,
however, is still difficult to assess in general.