Lecture 10: Control of enzyme activity Flashcards
What are the two main classes of inhibitor and how do they differ in their binding to an enzyme?
Irreversible inhibitors and reversible inhibitors
Irreversible inhibitors bind to the enzyme and permanently inactivates it, inhibitor reacts with a specific amino acid side chain, usually in the active site and forms a covalent bond
Reversible inhibitors bind to the enzyme but can be subsequently released (leaving enzyme in original condition)
What effect does a competitive inhibitor have on KM and Vmax in an enzyme catalysed reaction?
Competitive inhibitor competes directly with the substrate, for the active site
It will increase Km as more substrate is required to get
V = Vmax / 2
There is no change in Vmax as infinite [S] outcompetes the inhibitor
How can competitive inhibition of an enzyme be overcome ?
By increasing the substrate concentration, thus increasing the likelihood the enzyme will bind a substrate instead of an inhibitor and thus negate the effect of competitive inhibition.
Where on an enzyme does a competitive inhibitor bind?
It binds directly to the active site of the enzyme thus blocking normal function
What effect does a pure non-competitive inhibitor have on KM and Vmax in an enzyme catalysed reaction?
Binding changes the structure of the active site such that substrate still binds, but transition state stabilisation is no longer optimal
Km remains the same as the enzyme-substrate complex can still form with the same affinity, but the rate of conversion to product is reduced due to the inhibitor’s presence.
Vmax decreases as a non-competitive inhibitor binds to an allosteric site on the enzyme, not the active site, which affects the enzyme’s ability to catalyze the reaction regardless of the substrate concentration.
Where on an enzyme does a non-competitive inhibitor bind?
A non-competitive inhibitor binds to an allosteric site on the enzyme, not the active site
What is mixed non-competitive inhibition and what effect does it have on KM and Vmax in an enzyme catalysed reaction?
V max is decreased.
𝐾𝑚 increases
What are allosteric enzymes?
Allosteric enzymes are a class of enzymes that exhibit regulation by molecules that bind to sites other than the active site, known as allosteric sites. These enzymes have a more complex regulation mechanism compared to simple enzymes and often display more intricate kinetic behavior.
These regulations and factors such as cooperativity allow the enzyme to respond dynamically to changes in the cellular environment and is crucial for controlling various metabolic pathways.
Do allosteric enzymes follow the Michaelis-Menten model? Why/why not?
Allosteric enzymes do not follow the Michaelis-Menten model due to their cooperative binding properties, multiple conformational states, and regulation by allosteric effectors. These factors lead to more complex kinetics that are better described by models specifically designed for allosteric enzymes.
Briefly outline the features of the concerted model versus the sequential model.
Concerted Model (MWC Model):
- Conformational States: All subunits are in the same state (R or T).
- Transition: All-or-nothing transition between R (active) and T (inactive) states.
- Cooperativity: Simultaneous change in affinity for all subunits.
- Kinetics: Sigmoidal curve due to cooperative binding.
Sequential Model (KNF Model):
- Conformational States: Subunits independently transition states.
- Transition: Gradual, sequential changes in subunits upon substrate binding.
- Cooperativity: Binding changes one subunit and affects neighboring ones.
- Kinetics: Sigmoidal curve due to gradual cooperativity.
Briefly outline how the glycogen phosphorylase enzyme is
regulated
Glycogen Phosphorylase is fundamentally important in glucose metabolism
Has Feedforward activation - high AMP promotes enzyme activity - AMP activates glycogen phosphorylase by changing its conformation from the T-state to the R-state
Has Post translational modification regulation via Serine-phosphorylation (allosteric activation)
Has Feedback inhibition - increased Glucose-6-P can be stored and feedbacks to inhibit enzyme activity
Has non-competitive inhibition via Caffeine and Purines
What are some examples of allosteric enzymes and what types of pathways are they important in?
Haemoglobin - important in oxygen transport
Glycogen Phosphorylase - important in glucose metabolism
How does the presence of an allosteric inhibitor affect the
𝑉 vs [S] curve of a multimeric enzyme?
Inhibitor Stabilization:
Inhibitors stabilize the T (low affinity) state of the enzyme.
Curve Shape:
The V vs [S] curve becomes more sigmoidal.
Shift:
The curve shifts to the right.
Reason:
The enzyme relies more on cooperativity to increase substrate affinity, making the curve more sigmoidal.
How does the presence of an allosteric activator affect the 𝑉 vs [S] curve of a multimeric enzyme?
Activator Stabilization:
Activators stabilize the R (high affinity) state of the enzyme.
Curve Shape:
The 𝑉 vs [S] curve becomes more hyperbolic.
Shift:
The curve shifts to the left.
Reason: The enzyme has a high affinity for the substrate and is less dependent on cooperativity, resembling Michaelis-Menten behavior.
What are 2 examples of competitive inhibitors?
- Transition state analogues (e.g. Lipitor inhibits HMG-CoA reductase)
- Anastrozole - inhibits aromatase, preventing the conversion of testosterone into estradiol (can be used as treatment for some forms of breast cancer)
