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Chapter 8 key terms/concepts
Terms in this set (88)
An enzyme without its cofactor
The complete catalytically active enzyme.
Apoenzyme + cofactor = holoenzyme
Cofactors can be divided into two groups:
2. small organic molecules called coenzymes
Tightly bound coenzymes are called the prosthetic group
What does it mean to have a positive delta G?
an input of free energy is required to drive such a reaction, enderogonic
Delta G is independent of _______
the path of the transformation and provides no information about the rate of a reaction
True or false: the amount of product formed is the same whether or not the enzyme is present.
True, an enzyme cannot alter the equilibrium of a chemical reaction
Why does the rate of product formation level off with time?
The reaction has reached equilibrium
Substrate S is still being converted into product P, but P is being converted into S at a rate such that the amount of P present stays the same
What do enzyme do?
Enzymes accelerate the attainment of equilibrium but do not shift their positions.
The equilibrium position is a function only of the free - energy difference between reactants and products
Gibbs free energy
The difference in free energy between the transition state and the substrate
How do enzymes accelerate reactions?
By decreasing the activation energy. Because the activation energy is lower, more molecules have the energy required to reach the transition state
The specific region where substrates are bound in the enzyme - substrate (ES) complex
What is the evidence of an enzyme - substrate complex?
1. At a constant concentration of enzyme, the reaction rate increases with increasing substrate concentration until a maximal velocity is reached.
2.x- ray crystallography
3. The spectroscopic characteristic of many enzymes and substrates change on the formation of an ES complex
The residues that directly participate in the making and breaking of bonds.
What promotes the formation of the transition state?
The interaction of the enzyme and substrate at the active site
Generalizations of active sites
1. The active site is a 3D cleft, or crevice
2. the active site makes up a small part of the total volume of an enzyme
3. Active sites are unique microenvironments
4. Substrates are bound to enzymes by multiple weak attractions
5. The specificity of binding depends on the precisely defined arrangement of atoms in an active site
The active site of some enzymes assumes a shape that is complementary to that of the substrate only after the substrate has been bound.
The free energy that is released on binding. The maximal binding energy is released when the enzyme facilitates the formation of the transition state.
Most- stable interaction, least stable interaction
The most stable interaction (maximum binding energy) takes place between the enzyme and the transition state, the least stable is the transition state
The enzyme is still actively converting the substrate into product and vice versa but there is no net change in the concentration
The concentration of intermediates stays the same even if the concentrations of starting materials and products is changing
The maximal rate is attained when the catalytic sites on the enzyme are saturated with substrate
Is equal to the substrate concentration at which the reaction rate is half its maximal value.
Is the concentration of a substrate at which half the active sites are filled and provides a measure of the substrate concentration required for significant catalysis to take place
When is km equal to the dissociation constant of the ES complex?
When K2 is much smaller than K-1
When this condition is met, Km is a measure of the strength of the ES complex. High Km indicates weak binding. Low Km indicates strong binding
What does the maximal rate, Vmax, reveal?
The turnover number of an enzyme, which is the number of substrate molecules converted into product by an enzyme molecule in a unit time when the enzyme is fully saturated with substrate.
What does it mean when an enzyme has attained kinetic perfection?
Their catalytic velocity is restricted only by the rate at which they encounter substrate in the solution
For catalytically perfect enzymes, every encounter between enzyme and substrate is productive. In these cases, there may be attractive electrostatic forces on the enzyme that entice the substrate to the active site.
All substrates must bind to the enzyme before any product is released
A ternary complex of the enzyme and both substrates forms
Double - displacement (ping - pong) reactions
one or more products are released before all substrates bind the enzyme
What is the defining feature of double - displacement reactions?
The existence of a substituted enzyme intermediate, in which the enzyme is temporarily modified
what is an important group of enzymes that do not obey Michaelis - Menton kinetics?
Dissociates very slowly from its target enzyme because it has become tightly bound to the enzyme, either covalently or noncovalently
Characterized by a rapid dissociation of the enzyme - inhibitor complex
A form of reversible inhibition in which an enzyme can bind substrate (forming an ES complex) or inhibitor (EI) but not both.
Diminishes the rate of catalysis by reducing the proportion of enzyme molecules bound to a substrate
Can be relieved by increasing the substrate concentration
Distinguished by the fact that the inhibitor binds only to the enzyme - substrate complex.
Cannot be overcome by the addition of more substrate
the inhibitor and substrate can bind simultaneously to an enzyme molecule at different binding sites.
Can bind free enzyme or the enzyme - substrate complex.
Acts by decreasing the concentration of functional enzyme rather than by diminishing the proportion of enzyme molecules that are bound to substrate
Cannot be overcome by increasing the substrate concentration
Produced when a single inhibitor both hinders the binding of substrate and decreases the turnover number of the enzyme
Effects of competitive inhibitor
An enzyme will have the same Vmax as in the absence of an inhibitor. At sufficiently high concentration, virtually all the active sites are filled by substrate, and the enzyme is fully operative.
Km rises, takes more substrate to reach the same point
X shaped graph
Effects of uncompetitive inhibition
This enzyme - substrate inhibitor complex, ESI, does not go on to form any product. Because some unproductive ESI complex will always be present, Vmax will be lower in the presence of the inhibitor
Lowers value of Km because the inhibitor binds to ES to form ESI, depleting ES
Parallel shaped graph
Effects of noncompetitive inhibitor
The EI complex does not proceed to form product. The value of Vmax is decreased, whereas the value of Km is unchanged
V shaped graphy
In noncompetitive inhibitor why is Vmax lowered though Km remains unchanged?
The inhibitor lowers the concentration of functional enzyme
Covalently bond to the enzyme to provide an alternative and often complementary approach to determine functional groups. Divided into three categories
Group Specific Reagents
React with specific side chains of amino acids
Or reactive substrate analogy, are molecules that are structurally similar to the substrate for an enzyme and that covalently bind to active - site residues.
Or mechanism - based inhibitors, are modified substrates that provide the most specific means for modifying an enzyme's active site.
The inhibitor binds to the enzyme as a substrate and is initially processed by the normal catalytic mechanism.
The mechanism of catalysis then generates a chemically reactive intermediate that inactivates the enzyme through covalent modification
How do enzymes affect the transition state?
Enzymes accelerate reactions by stabilizing the transition state
What is methotrexate a competitve inhibitor of?
Transition state analogs
Since enzymes work by stabilizing the transition state, they bind most strongly to transition state analogs. These ts analogs can then help us understand the mechanism of the enzyme
Develope an antibody that selectively binds the transition state of a reaction. The antibody then becomes a catalyst
How does penicillin operate?
Penicillin irreversibly inactivates enzymes involved in cell wall synthesis
How much product is created in a reaction depends on what aspects?
Depends on the energetics of the chemical reaction and environmental conditions where the reaction takes place (temperature, S, P)
How do enzymes attract substrates?
Enzymes have a "pocket" that attracts its substrates through chemical interactions, and brings them into the transition state. A chemical shift in the enzyme takes place once they do allowing the reaction to take place, and the products are released, freeing the enzyme to accept more reactants.
What does K1 represent in the Michaelis - Menton equation?
The rate at which (ES) is formed from (E) and (S)
What does k-1 represent in the Michaelis - Menton equation?
The dissociation of (ES) into (E) and (S) again
What does K2 or kcat represent in the Michaelis - Menton equation?
The rate at which (ES) actually goes onto form (P) and release (E) again
Kcat / Km
Is a measure of enzyme efficiency; the larger it is, the faster the rate is for the chemical reaction your're measuring it for
What does it mean when the concentration of the substrate is equal to Km?
your rate is half its maximum speed
Suppose that a mutant enzyme binds a substrate 100 times as tightly as does the native enzyme. What is the effect of this mutation on catalytic rate if the binding of the transition state is unaffected?
The substrate would be so stable that it would be less likely to change to the transition state because the energy of activation would be effectively increased.
The reaction is slowed
For a one - substrate, enzyme catalyzed reaction, double - reciprocal plots were determined for three different enzyme concentrations. What would you expect the graph to look like and why?
The graph would look like the same linear line but at different slopes. This is because if the total amount of enzyme is increased, Vmax will increase. Vmax is independent of the substrate concentration
What are some assumptions that are made if given the Michaels - Mention equation?
V = (ES)k2 how fast you get product
Steady State approximation, ES complex is at "steady state" the amount of ES is not changing because the rate of formation of ES is the same as the rate of break down of ES
What does the Michaels - Menton equation tell us?
The velocity of enzyme reaction, the speed of product formation
Where does competitive inhibitors bind? uncompetitive? noncompetitive?
competitive = in active site
uncompetitive = Es complex
noncompetitive = Binds to E or ES, but not in active site
What does a low Km indicate? higher km?
lower km indicates tighter binding (further left on x axis), where higher km indicates weaker binding (further right on x axis)
What is an example of an uncompetitive inhibitor?
Summary of Reversible inhibitors
Competitive: Km higher, V max Same
Uncompetitive: Km Lowered, Vmax Lowered
Noncompetitive: Km same, Vmax Lowered
Which of the statements is true regarding the role an enzyme plays in catalysis?
An enzyme decreases the activation energy
The commonly used derivation of the Michaelis-Menten uses the "steady state assumption". Which of the following does NOT describe this assumption?
The concentrations of enzyme and substrate do not change significantly during the course of the reaction.
Vmax, the maximum velocity, of an enzyme - catalyzed reaction is
the rate observed when all enzyme active sites are saturated with substrate
In a double - reciprocal or Lineweaver - Burke plot, the Km is found from
x - intercept = -1/km
TPCK, a molecule with larger hydrophobic groups, inactives chymotrypsin but not trypsin because:
it looks like the substrate for chymotrypsin (but not trypsin) and thus can bind in its active site and modify His - 57
Which of the following is Not true of a competitive inhibitor?
It irreversibly inhibits the enzyme by chemically modifying a group at the active site.
If an enzyme inhibitor decreases the apparent km and reduces the vmax as well, the nit can be classified as a (N)
When k-1 > k2 (that is, when the rate constant for dissociation of the enzyme substrate complex is greater than the rate constant for conversion to product), the Km is most analogous to
A reaction has an equilibrium constant, Keq, of 50. When performed in the presence of an appropriate enzyme, the forward rate constant is increased 20-fold. What will happen to the reverse rate constant?
It will increase 20 - fold
To obtain k2, the turnover number of an enzyme, one must
divide vmax by the total enzyme concentration
The rate constant kcat/ Km is a measure of catalytic efficiency because... ?
It takes into account both the rate of catalysis with a particular substrate (kcat) and the strength of the enzyme - substrate interaction (Km).
What does a competitive inhibitor resemble?
The competitive inhibitor often resembles the substrate and binds to the active site of the enzyme
What is the net effect of a noncompetitive inhibitor that acts by decreasing the concentration of functional enzyme rather than by diminishing the proportion of enzyme molecules that are bound to substrate.
The net effect is to decrease the turnover number
What happens to the delta G when there is a 10 fold change in the equilibrium constant?
The delta G changes by 5.69 kj or (1.36 kcal)
what is the difference between a rate constant and an equilibrium constant?
rate constant only "look" at a reaction in a single direction, whereas equilibrium constant "look" at both directions.
Why do allosteric enzymes not work with the Michaelis - Menten Equation?
Because the Michaelis - Menten equation assumes a single active site and a single substrate, where an allosteric enzyme such as Hb have multiple interacting sites.
If a low Km indicates tighter binding how would it be possible to achieve tighter binding and a lower km?
The equation for km =( K-1 + K2 / K1) therefore to get a lower km one would have to increase the K1
Type of reaction: Oxidation - reduction
Example: Lactate dehydrogenase
Type of reaction: Group transfer
Example: Nucleoside monophosphate kinase (NMP kinase)
Type of reaction: Hydrolysis
Type of reaction: Addition or removal of groups to form double bonds.
Type of reaction: Isomerization
Example: Triose phosphate isomerase
Type of reaction: Ligation of two substrates at the expense of ATP hydrolysis
Example: Aminoacyl - tRNA synthetase
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