Chpt 7-Models for Enzyme Kinetics and classes of enzyme inhibitor drugs Flashcards
Enzyme kinetics help to do what?
- Understand general cell metabolism
- Understand inhibitor drug action
- Design therapeutic agents
Kinetic assays with enzymes are widely used in drug _____________.
discovery and development.
What tests for the presence or absence of specific enzymes employ kinetics?
Clinical diagnostic
Higher the [S] greater the [P] formation.
However, the ______ formation is indirectly
dependent on [E.S].
Product [P]
What is explained in Michaelis-Menten Model?
- Enzyme contacts substrate by DIFFUSION and forms the E∙S reaction intermediate.
- Then E∙S passes over an activation barrier to yield product P
- Or E∙S could fall apart and the enzyme and substrate diffuse away from each other.
Enzyme contacts substrate by _________ and forms the E∙S reaction intermediate.
diffusion
many enzyme-catalyzed reactions are reversible or irreversible?
Irreversible
An ACTIVATED complex (E.S)‡ has no _______.
Stability. (unstable). It is the state at the highest free energy level (top) of the reaction progress diagram.
How long does the activated complex last?
It lasts for only one or a few molecular vibration periods; about 10-15 sec.
The (E•S) complex is generally much ______ compared to activated E•S and may last for quite some time.
Stable
k−1 is the rate constant for the ___________. A uni-molecular reaction step.
dissociation of E∙S
What is the rate constant for forming E∙S?
k+1. A bimolecular reaction step.
k+2 is the rate constant for the breakdown of E∙S to form E + P. Also a ______ reaction step.
unimolecular
This number defines the maximum number of substrate molecules converted
k+2 or kcat
Apparent second-order kinetics gradually switches over to first-order kinetics when?
An unusual feature in enzyme kinetics: as the concentration of substrate increases
When [S] is _____ by comparison to Km we have mostly free enzyme in the system
low
When [S] is high, we have mostly enzyme with substrate in the ________
active site
What is the first regime of substrate kinetics?
- Mostly saturated enzyme
- Depends only on [E0]
- Maximal rate of reaction
The higher the Km of an enzyme, the ________ its affinity for its substrate.
LOWER
Mostly free enzyme, Depends on both [E] and [S], k+2 / KM appears to be an effective second-order rate constant defines what?
Second-order kinetics : [S] «_space;Km
The traditional unit (U) of enzyme activity is the amount of enzyme that can convert one micromole of substrate into products in one minute.
units of enzyme activity
SI unit of activity is the katal (symbol kat), the amount of activity that converts one mole of substrate into products in one second.
units of enzyme activity
How does many drugs function to inhibit key enzymes?
By blocking their action, a therapeutic effect is produced
drugs “compete” with substrate, for access to the enzyme called
competitive inhibition
Competitive inhibition effects the catalytic process. T or F?
False. A competitive inhibitor has no effect on the catalytic step; kcat or k+2 are the same.
Formation of the E∙S complex is reduced while a new type of complex, E∙I, is formed.
This occurs in competitive inhibition
The apparent Michaelis constant is numerically ________ than Km and it increases as the concentration of inhibitor rises.
Larger. when referring to competitive inhibition
Can a higher concentration of substrate restore the original rate of reaction (overcome the inhibition) regarding competitive inhibition?
Yes
The maximum velocity is still the same; Vmax does not change. T or F
True
The E∙I and E∙S∙I complexes are both active or inactive for mixed and competitive inhibition?
Inactive
For noncompetitive inhibition the inhibitor has the ________ affinity for either E or the E∙S complex
Same
The irreversible inhibition of an enzyme entails ______ attachment of the inhibitor to the enzyme, or some covalent modification, involving key residues of the enzyme, by the inhibitor.
- Covalent
Many drugs that target enzymes are what kind of inhibitors?
Irreversible inhibitors of those enzymes like penicillin and aspirin
Active-site directed irreversible inhibition are designed to do what?
These inhibitors are designed to attach covalently in or near the enzyme’s active site
Suicide inhibitors are another class of _______ inhibitors
irreversible
What kind of inhibition is describe as the inhibitor being unreactive until the enzyme attempts to use it as a substrate?
Suicide inhibitors
Upon binding and undergoing few chemical changes the molecule becomes highly reactive and binds irreversibly with the enzyme. Thus the enzyme commits “suicide” by converting the compound.
Suicide inhibitors for example Penicillin, physostigmine, 5-fluorouracil
Many enzymes have _______ groups that are needed either for catalytic activity, ___________ , or for structural reasons.
- Sulfhydryl
2. Binding cofactors
These sulfhydryl groups can form tight bonds with heavy metals such as mercury, lead, silver, even iron and copper.
Heavy Metal Toxicity
Once the heavy metal atom is bound, the sulfhydryl cannot function in catalysis
Heavy Metal Toxicity
This defines Enzymes as drug targets
Pick a key enzyme, inhibit it, reduce the concentration of the metabolites and/or buildup the concentration of substrate(s), and so get a clinically useful response.
Give examples of Enzymes as drug targets
- Statins inhibit HMGCoA reductase, leading to lower cholesterol levels.
- Gleevec inhibits a protein kinase and blocks proliferation of cancer cells.
- Aspirin inhibits a cyclo-oxygenase and reduces inflammation.
Combine two inhibitors that target the same (or a related) pathway to get an enhanced response, greater than what might be achieved with either inhibitor alone. This is the definition of what?
Combination chemotherapy
Give an example of combination therapy
Example: Co-trimoxazole is a very effective antibacterial combination of trimethoprim and sulfamethoxazole
What are the four basic strategies of inhibitor design?
- Mimic the substrate;
- Mimic the transition state;
- Irreversibly react with a key residue;
- Change the enzyme’s conformation or state of assembly
Substrate mimics are usually ______ inhibitors
competitive
use compounds that have much but not all of the chemical features of the substrate, so that they fit into the active site and are held relatively tightly, but do not themselves react and do not let substrate react.
Substrate mimics
Give an example of a substrate mimic
Examples: Ibuprofen and related compounds occupy the active site to inhibit prostaglandin synthase (COX) enzyme, and act as anti-inflammatory drugs
Compounds can be made that ________ the transition state of substrate but that cannot go on to complete the reaction.
Resemble
Transition state compounds
These compounds should bind more tightly to the enzyme than does the natural substrate
Irreversible Inhibition
Aspirin acetylates a key serine residue in the active site of cyclo-oxygenase (prostaglandin synthase)
Change the Enzyme’s Conformation or State of Assembly
The inhibitor might then interfere with binding of substrate in the correct conformation, or with catalysis in some way.
Use an uncompetitive inhibitor that might bind to a different site on the enzyme, away from the active site.
Change the Enzyme’s Conformation or State of Assembly
Another possibility is that the inhibitor could block the proper association of enzyme subunits.
Such inhibitors are much harder to discover and develop, and are not very common
Change the Enzyme’s Conformation or State of Assembly
