Transcript Center for Structural Biology

Enzymes: Protein Catalysts
• Increase rates of reaction, but not consumed.
• Enable reactions to occur under mild conditions: e.g.
temperature, pH.
• High reaction specificity: no side products.
• Activity of enzymes can be regulated.
 Substrate
 Availability of enzyme (expression, localization)
 Reversible covalent modification
Allosteric control (other proteins or co-factors)
Enzyme Cofactors
1. Simple Enzymes: only protein chain(s)
2. Complex Enzymes: require additional molecules (cofactors) for functioning
 Co-enzyme: non-covalently bound
 Prosthetic Group: covalently bound to protein
Prosthetic groups/co-enzymes usually VITAMINS!
Vitamin deficiency diseases: malfunction of enzymes
Enzyme Classes/Reactions
1. Oxidoreductases. Act on many chemical groupings to add or
remove hydrogen atoms.
2. Transferases. Transfer functional groups between donor and
acceptor molecules (e.g. kinases that transfer phosphate
from ATP to other molecules).
3. Hydrolases. Add water across a bond, hydrolyzing it.
4. Lyases. Add H2O, NH2, or CO2 across double bonds, or
remove these elements to produce double bonds.
5. Isomerases. Carry out stereochemical, geometric and other
types of isomerization.
6. Ligases. Catalyze reactions in which two chemical groups
are joined (ligated) with the use of energy from ATP.
Catalytic Mechanisms
1. Bond Strain: Strains substrate bonds, which facilitates
attaining the transition state.
2. Proximity and Orientation: Binding brings molecules
into proximity and helps to properly orients reactive
groups.
3. Acid/Base Catalysis: Required catalytic proton donors
(acids) or acceptors (bases) are supplied by catalyst.
4. Covalent Catalysis: The reaction is facilitated by
formation of a covalent intermediate between the
enzyme (or coenzyme) and the substrate.
Catalytic Mechanisms (cont.)
5. Metal Ions (metalloenzymes): Orients substrates +
stabilizes charge in the transition state + supplies or
captures electrons during the course of the reaction.
6. Electrostatic: Exclusion of solvent changes
environment to facilitate reactions + stabilization of
charge in the transition state.
7. Preferential Binding of the Transition State: Binding
of the transition state is PREFERRED over the
reactants and products, greatly stabilizing this
species at the critical moment it is formed.
Enzymes: Protein Catalysts
• Catalysts: increase rate of reaction, but not consumed.
• Enzymes bind substrates to increase the rate of a
biochemical reaction that converts the substrate
(reactant) into a desired product.
• Reactions occur once sufficient energy has been
supplied to overcome the energy barrier that prevents
the reaction from occurring spontaneously.
• Each reaction has a transition state where the substrate
is in an unstable, short-lived chemical/structural state.
A-H + B  [ AHB ]  A + H-B
Reaction Coordinate Diagrams
• The progress of the reaction can be viewed in terms of
the energy of the system: reaction coordinate. [Fig. 11-3]
• The transition state is the highest point on the reaction
coordinate diagram.
• The height of the energy barrier is the Free Energy of
Activation and is symbolized by DG‡.
• Reactions can go forward and backward. The height of
the energy barrier to go backwards may be higher than
to go forward, because the product may be more stable
than the substrate. The difference in energy between
substrate and product is the Free Energy of Reaction.
Multi-Step Reactions
• A reaction may have more than one step: S  I  P.
• Two step reaction have two transition states.
[Fig. 11-4]
• If the energy barrier is higher for one step than the
other, than the rate of this step will be slower.
• In multi-step reactions, the step with the highest
transition state free energy (the highest point on the
reaction coordinate diagram) is the Rate Determining
Step of the reaction.
• The overall rate of the reaction, kinetics, can only be as
fast as the slowest step.
The Catalytic Effect of Enzymes
• Enzymes act by lowering the free energy of the
transition state, thereby reducing the free energy of
activation (DG‡).
[Fig. 11-5]
• The catalytic efficiency (DDG‡cat) is the difference in DG‡
for the catalyzed reaction versus the uncatalyzed
reaction.
• The catalytic efficiency can be viewed in terms of the
kinetic parameters: rate enhancement for the reaction.
• Enzymes allow huge enhancements of rates, in many
cases, enabling reactions to occur that would almost
never occur spontaneously because DG‡ is so high.
Enzymes Affect Rates, Not Outcome
• The lowering of the free energy barrier and increase in
rate is equal for the forward and the reverse reactions.
• Ultimately, the reaction will settle to equilibrium. The
time required to come to equilibrium depends on the
rates of the forward and reverse reactions. Enzymes
facilitate this process.
• Enzymes increase the kinetic parameter velocity with
which products are produced from reactants. This is
equally true for the forward and reverse reactions.
• At equilibrium, the preference for substrate versus
product is determined by the difference in energy
between substrate and product- NOT BY THE RATES.
Simple Kinetics
S ------> P
Reaction Velocity: the instantaneous rate (k) at which
product is produced (or substrate disappears) mulitplied
by the amount of substrate present: v = k[S]
The rate of production of product = rate of consumption
of substrate.
The rate constant for the forward reaction is defined as
kf or k+1 and for the reverse reaction as kr or k-1:
The velocity of the forward reaction is: vforward = k+1[S]
The velocity of the reverse reaction is: vreverse = k-1[P]
Kinetics at Equilibrium
At equilibrium, the velocity of the forward reaction is
equal to the reverse reaction k+1[S] = k-1[P], so there is
no overall change in the distribution of S and P.
An equilibrium constant of the reaction can be
defined:
Keq = [P]/[S] = k+1/k-1
This equation demonstrates that at equilibrium, the
concentration of S and P reflects the relative stability
(free energy difference) between S and P.