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Concepts to understand
Enzyme terminology
____: The molecule on which an enzyme acts.
Catalytic center or Active site: The specific site where the substrate binds to an enzyme.
Transition state: Intermediate state formed by enzyme and substrate
Enzyme-substrate complex: is formed in the transition state
___ ___: An energy barrier that reacting molecules must overcome to form products.
___: Molecules or ions that are essential for the catalytic activity, includes metals or small organic molecules
Km = measure of E-S affinity
km = __ at Vmax/2
Reaction reaches maximum rate at lower [S] when Km is ___
Two ways to change Vmax :
1) K2
2) [E]
Enzyme Regulation
In health:
• Enzymes must be tightly ___ to do the work only where and when needed and not to cause damage.
In disease:
• Enzymes can be drugs (e.g., digestive enzymes)
• Enzymes can be targeted by drugs,
for example by regulating their activity.
*Many drugs are enzyme ____.
Enzyme inhibition
-Inhibition is one of the fastest regulation mechanisms
-An inhibitor can bind to an enzyme to __ down the reaction rate V.
The binding can be:
A) reversible (reversible inhibition). Weak non-covalent bonds are formed and easily broken.
B) irreversible (irreversible inhibition). Often covalent bonds are formed. Non-covalent bonds can form as well. The enzyme often becomes ___.
Reversible inhibition:
There are three major inhibition mechanisms that demonstrate
different enzyme kinetics:
1. Competitive inhibition (la. classical and lb. non-classical)
2. Non-competitive inhibition
3. Uncompetitive inhibition
1. Competitive inhibition
What happens to Vmax?
Km?
2. Non-competitive inhibition
Vmax?
km?
3.Uncompetitive inhibition
Vmax?
km?
Competitive Inhibition
a) Classical competitive inhibition
-Inhibitor resembles substrate
-inhibitor ___ the ES time and therefore,
also __ affinity
**This is why Km ___
Competitive Inhibition
b) Non-classical competitive
-Inhibitor does __ have to resemble the substrate
-Allosteric inhibition
-They are still competing in a similar way as in the case of classical inhibition (Km still increases & Vmax remains unchanged).
Uncompetitive inhibition
Inhibitor can only bind if the substrate is ___ to the enzyme
S become ___ regulator for I
Vmax ___ & Km ___ so the slope remains the same
Noncompetitive inhibition
-Inhibitor does not have to resemble substrate
-Inhibitor binds __ of S-binding
-Allosteric inhibition
-Vmax ___
-Km ___
Mixed Inhibition
Enzymes cannot only be inhibited allosterically but also ____
Example: Phosphofructokinase
Example: Phosphofructokinase & feedback loop
• When ATP levels drop low, [ADP] and, especially, [AMP] increases
-Glycolysis needs to produce more ATP
• AMP and ADP allosterically activate _____
Phosphofructokinase-1 is a tetramer, 4 identical subunits
-Active site is physically ___from regulatory site
If glycolysis stops at the 10th stage, phosphoenolpyruvate (PEP) will accumulate.
PEP is allosteric ___ for Phosphofructokinase (PFK).
PEP binding to a subunit of PFK, switches it to T-conformation.
PEP inhibition is an example of a negative feedback loop:
Up-stream reaction is regulated by ___-stream product
Phosphofructokinase tetramer may behave in a ___way, similar to hemoglobin.
R-conformation of one subunit forces other units to switch from T to R.
Kinetics may be quite complex.
R - Relaxed, ___-affinity
T — Tense, ___-affinity
Enzyme kinetics for a simplified case:
- Substrate and activator only bind enzyme in the __ state
- Inhibitor only binds enzyme in the ___ state
- There is a fast interconversion between R and T
Regulation by covalent modification
One enzyme can regulate another using (de)phosphorylation
Usually ___ than allosteric regulation
-It is reversible
