MODULE 7: Chapter 7.5 Flashcards
What are the two primary mechanisms of enzyme regulation?
- Bioavailability 2. Control of catalytic efficiency through protein modification
Bioavailability pertains to the amount of enzyme in different tissues and cellular compartments, while catalytic efficiency is influenced by covalent and noncovalent modifications.
What can positive regulatory mechanisms lead to in enzyme activity?
An increase in enzyme activity
This can result from increased enzyme synthesis or enhanced catalytic efficiency due to conformational changes.
What are the biochemical processes affecting enzyme bioavailability?
- RNA synthesis (gene transcription)
- RNA processing
- Protein synthesis
- Protein degradation
- Protein targeting
These processes influence the amount of enzyme available for catalytic activity.
What are the three primary mechanisms that affect catalytic efficiency?
- Binding of regulatory molecules
- Covalent modification
- Proteolytic processing
These mechanisms help control enzyme activity and responsiveness.
What is enzyme inhibition?
A regulatory mechanism to control enzyme activity
Enzyme inhibitors can also be used in research and therapy to affect enzyme function.
What are the two types of enzyme inhibition?
- Reversible inhibition
- Irreversible inhibition
Reversible inhibition involves noncovalent binding, while irreversible inhibition involves covalent bonds.
How can reversible inhibitors be affected by enzyme dilution?
The effect of reversible inhibitors can be decreased by diluting the enzyme reaction
This is because the noncovalent interactions can be disrupted.
What is malonate’s role in enzyme inhibition?
Malonate is a reversible inhibitor of succinate dehydrogenase
It competes with succinate for binding to the active site, inhibiting the enzyme’s function.
What characterizes irreversible inhibitors?
They form a tight complex with the enzyme, effectively ‘killing’ it
This occurs through covalent bonds or very strong noncovalent interactions.
What is a suicide inhibitor?
A type of irreversible inhibitor that reacts with an enzyme during catalysis but does not complete the reaction
It forms a covalent bond and remains irreversibly bound to the enzyme.
What are the three classes of reversible inhibitors?
- Competitive inhibitors
- Uncompetitive inhibitors
- Mixed inhibitors
These classes differ in their binding mechanisms and effects on enzyme kinetics.
What defines a competitive inhibitor?
It inhibits substrate binding at the active site
Competitive inhibitors can bind directly to the active site or partially obstruct it.
What happens to the apparent Km in the presence of a competitive inhibitor?
The apparent Km increases
This reflects the requirement of higher substrate concentration to reach vmax.
What is the effect of uncompetitive inhibitors on Km and vmax?
Both Km and vmax decrease
The net effect is a constant slope of Km/vmax in a Lineweaver–Burk plot.
How do mixed inhibitors function?
They bind to both the enzyme and the enzyme–substrate complex
This results in a decreased vmax and forms unproductive EI or ESI complexes.
What is the role of structure-based drug design in enzyme inhibition?
It uses knowledge of an enzyme’s structure to create inhibitors that fit the active site
This strategy is employed in the development of drugs targeting specific enzymes.
What is the equilibrium dissociation constant KI?
It describes the affinity of an enzyme for an inhibitor
KI specifically refers to the equilibrium dissociation constant for an enzyme–inhibitor complex.
How do competitive inhibitors affect enzyme kinetics in a Lineweaver–Burk plot?
Vmax remains unaffected, but the apparent Km increases
This is evident in the shifting of the Lineweaver–Burk plot.
What is the effect of increasing substrate concentration in the presence of a competitive inhibitor?
The effects of the inhibitor can be overcome
High substrate concentrations can outcompete the inhibitor for binding to the active site.
What is mixed inhibition?
A mixture of competitive and uncompetitive inhibition, where an unproductive EI or ESI complex is formed, decreasing the enzyme’s catalytic activity.
Mixed inhibition results in decreased vmax and may increase or decrease Km depending on the relative KI and KI’ values.
What happens to vmax in the presence of a mixed inhibitor?
Decreased.
The effect is due to some enzyme being bound to the inhibitor, making less enzyme available.
What is noncompetitive inhibition?
A special case of mixed inhibition where the inhibitor has equal affinity for both the free enzyme and the ES complex (KI = KI’).
Noncompetitive inhibitors bind away from the active site and do not compete with the substrate.
What effect does noncompetitive inhibition have on Km?
Unchanged.
Both the free enzyme and the enzyme-inhibitor complex can bind to substrate.
How does feedback inhibition work?
The end product of a metabolic pathway inhibits the first enzyme in the pathway when it accumulates.
This mechanism helps regulate metabolic pathways efficiently.
What is the role of aspartate transcarbamoylase (ATCase) in metabolic regulation?
It is a key regulated enzyme in the pyrimidine biosynthetic pathway, catalyzing the formation of N-carbamoyl-L-aspartate.
ATCase is regulated by feedback inhibition from CTP and activation by ATP.
What distinguishes allosteric regulation from standard enzyme kinetics?
Allosteric regulation allows for a higher degree of control and does not follow Michaelis–Menten kinetics.
Cooperative binding leads to a sigmoidal activity curve.
What are homotropic allosteric activators?
Substrates that affect the binding of the same molecules at other active sites.
In the case of ATCase, carbamoyl phosphate and aspartate are homotropic activators.
What are heterotropic allosteric regulators?
Molecules that bind to regulatory sites and affect the binding of different substrates at other sites.
CTP and ATP are examples of heterotropic regulators for ATCase.
What is the impact of phosphorylation on enzyme activity?
Phosphorylation can activate or inactivate enzymes by altering their structure and active site chemistry.
This is a common form of covalent modification in enzyme regulation.
What is the role of glycogen phosphorylase in glucose metabolism?
It degrades glycogen to generate glucose for energy conversion in glycolysis.
The enzyme is active in the phosphorylated R-state and inactive in the unphosphorylated T-state.
What is the significance of the regulatory site in glycogen phosphorylase?
It is not directly at the active site but affects activity through structural changes upon phosphorylation.
Phosphorylation shifts the equilibrium toward the active R-state.
What is adenylylation in the context of enzyme regulation?
A covalent modification that inactivates certain enzymes, such as E. coli glutamine synthetase, by adding nucleoside monophosphate groups.
This modification can shift the enzyme between active and inactive states.
What is the T-state and R-state conformation in enzymes?
T-state is the inactive form, while R-state is the active form of the enzyme.
The transition between these states involves significant conformational changes.
What is the role of glutamine synthetase in bacterial nitrogen metabolism?
It catalyzes an ATP-dependent reaction that incorporates free NH4⁺ into the amino acid pool by converting glutamate to glutamine.
What conformational states does adenylylated glutamine synthetase exist in?
Inactive T-state conformation and active R-state conformation.
What enzyme catalyzes the adenylylation and deadenylylation of glutamine synthetase in E. coli?
Glutamine synthetase adenylyltransferase.
How is the activity of glutamine synthetase adenylyltransferase regulated?
Through uridylylation of Tyr51 by uridylyltransferase.
Under what conditions is the uridylylation activity of uridylyltransferase stimulated?
Under conditions of high ATP and elevated levels of α-ketoglutarate.
What happens when Pi and glutamine levels are elevated in the cell?
The deuridylylation activity of uridylyltransferase is stimulated, activating adenylation activity.
What is a zymogen?
An inactive enzyme precursor that is activated by a proteolytic cleavage reaction.
What is pepsinogen and how is it activated?
Pepsinogen is a zymogen that catalyzes an autocleavage reaction to generate the active enzyme pepsin.
What is the pH environment that stimulates the autocleavage of pepsinogen?
pH 1.5.
What is the process by which chymotrypsinogen is converted to its active form?
Proteolytic cleavage events that result in the formation of α-chymotrypsin.
Which enzyme initiates the cleavage of chymotrypsinogen?
Trypsin.
What type of regulation does proteolytic cleavage represent?
Irreversible regulation.
What three primary mechanisms regulate enzyme activity in the cell?
- Competitive inhibition
- Allosteric regulation
- Reversible covalent modification.
What is competitive inhibition?
A mechanism where the product of a reaction competes for substrate binding.
What is negative allosteric regulation?
A mechanism where a metabolite acts as an allosteric effector molecule, altering enzyme activity.
What are kinases and phosphatases?
- Kinases: Enzymes that phosphorylate
- Phosphatases: Enzymes that dephosphorylate.
What is feedback inhibition?
An enzyme regulatory mechanism where the end product of a pathway functions as an inhibitor of the first enzyme in the pathway.
What is irreversible inhibition?
An enzyme regulatory mechanism in which an inhibitory molecule forms a covalent bond with catalytic groups in the enzyme active site.
What is a suicide inhibitor?
An inhibitor that initiates the enzyme reaction mechanism but does not complete it, remaining covalently bound to the active site.
What defines a competitive inhibitor?
A molecule that inhibits substrate binding at the active site.
What defines an uncompetitive inhibitor?
A molecule that binds to enzyme–substrate complexes and alters the active site conformation.
What is a mixed inhibitor?
A molecule that binds to sites distinct from the enzyme active site but can bind to both the enzyme and the enzyme–substrate complex.
What is noncompetitive inhibition?
A special case of mixed inhibition where a molecule has equal affinity for both the free enzyme and the enzyme–substrate complex.
What is adenylylation?
A covalent modification of an enzyme by addition of AMP that regulates catalytic activity.
What is uridylylation?
A process that regulates enzyme activity by controlling adenylylating and deadenylylating activity.
