Enzyme Activity Flashcards
Give the general equation for a reversible reaction
Substrate ↔ products
What must be true if a reaction is going to occur?
There must be a change in energy
Why must there be a change in energy for a reaction to occur?
Because of the making and breaking of bonds
What is important for overcoming energy barriers?
Enzymes
Why are enzymes important for overcoming energy barriers?
They produce transition states
What is the transition state?
The high energy intermediate that lies between substrate and products
What is the activation energy?
The minimum energy that substrate must have to allow for the reaction
What must be done to increase the rate of reaction?
The energy barrier must be overcome
How can the rate of reaction be increased?
- Temperature
- Concentration
- Enzymes
How does temperature increase rate of reaction?
It increases the number of molecules with the activation energy
How does concentration increase in the rate of reaction?
It increases the chance of molecular conditions
What are the limitations of changing the rate of reaction using temperature and concentration?
They can’t really be changed in biological systems
What are enzymes?
Biological catalysts that increase the rate of reaction by lowering the activation energy
What do enzymes facilitate?
The formation of the transition state
Why is rate of reaction important?
Because some reactions are very slow, so even if it’s a favourable reaction, it may not occur
What are the features of enzymes?
- Highly specific
- Unchanged after reaction
What is the result of enzymes being highly specific?
They normally only work for 1 or 2 reactions
Do enzymes change at all?
They may be changed during the reaction
What do enzymes do to rate of reaction?
Increase it
What kind of molecules are enzymes?
Proteins (with one or two exceptions)
What may enzymes require?
Associated cofactors
What are cofactors?
Other things that allow enzymes to work
Why are enzymes relevant in medicine?
- Inheritable genetic disorders can be caused by changes in an enzyme
- Overactive enzymes can cause disease
- Measurement of enzyme activity for diagnosis
- Inhibition of enzymes for drugs
How can genetic disorders cause problems with enzymes?
Mutations can destroy or slow an enzyme
Give an example of a disease caused by an overactive enzyme?
Some cancers
What is the active site?
The place where substrates bind, and where chemical reactions occur
How much of the enzyme does the active site occupy?
Only a small part; 5-6 amino acids
What is the purpose of the rest of the enzyme?
Acts as a scaffold for the active sit
What is the active site formed from?
Amino acids from different parts of the primary sequence
How is it possible that the active site amino acids come from different parts of the amino acid sequence?
When the protein is in it’s 3D confirmation, the amino acids are bought close together
Where do amino acids usually sit?
In clefts or crevices- not usually on the surface
What does the creft/crevice in which the active site usually do?
Excludes water
Why does the cleft/crevice the active site sits in usually exclude water?
Because water is so concentrated that it can interfere with the reaction
What shape is the active site?
One that is complementary to the substrate
What does the binding of the substrate to the active site induce?
A change in confirmation
What is the theory that the binding of substrate to the active site causes a change in confirmation called?
The induced fit hypothesis
When does the active site form a completely complementary shape to the substrate?
After the substrate has binds
What happens as the substrate binds?
The enzyme changes shape to accommodate the substrate
How is the substrate bound to the active site?
By multiple weak bonds
What type of bonds bind the active site to the substrate?
All non-covalent
Why must the binding of active site to substrate not be too tight?
Otherwise the product would not be released
What is the importance of the non-covalent interactions binding the substrate to the active site?
They hold it in exactly the right conformation
How do enzymes work?
By stabilising the transition state
What happens as the substrate levels decrease?
Product levels increase
What is V 0 ?
The initial rate of reaction
Why do we use V 0 ?
Because when measuring rate of reaction, we must know the concentration of the substrate, and the only time we know this is right at the start
What do different enzymes have?
Different optimum conditions
What do optimum conditions consist of?
NAME?
What often happens as an enzyme catalysed reaction reaches maximum velocity?
The plot of reaction rate as a function of substrate concentration has the shape of a rectangular hyperbola
What does the Michaelis-Menten model for enzyme catalysis propose?
That a specific complex between enzyme and substrate is a necessary intermediate in catalysis
Give the chemical equation for the Michaelis-Menten model
E+S ↔ ES → E+P
What is the Michaelis-Menten equation?
V 0 = Vmax[S] / Km + [S]
Where Km = Michaelis constant
and V max = maximal velocity (mol/min)
Assuming an infinite [substrate]
What does the Michaelis-Menten equation predict?
A plot of V 0 versus [S] will be a rectangular hyperbola
What does the Michaelis-Menten equation tell us?
That rate is dependent on concentration
Do all enzymes obey the Michaelis-Menten model?
No
Which enzymes often don’t follow the Michaelis-Menten model?
Regulated enzymes that show co-operativity
What is Vmax?
The maximal rate when all enzymes active site are saturated with substrate
What does Vmax vary depending on?
Concentration of enzyme being measured
What is Km?
Substrate concentration that gives half maximal velocity
What do Km values give?
A measure of affinity of an enzyme to its substrate
What does a low Km mean?
High affinity for substrate
What does a high Km mean?
Low affinity for substrate
What is 1 unit?
The amount of enzyme that can catalyse 1 micromole of product per minute
What is the Lineweaver-Burk plot?
1/V 0 = Km/Vmax . 1/[S] + 1/Vmax
How can Km be determined from the Lineweaver-Burk plot?
It is the negative reciprocal of where the plot meets the x-axis
How can Vmax be determined from the Lineweaver-Burk plot?
It is the reciprocal of where the plot meets the y axis
What are enzyme inhibitors?
Molecules that slow down or prevent enzyme reactions
What do enzyme inhibitors include?
Many drugs
Are enzyme inhibitors reversible or irreversible?
Can be either
What happens when enzyme inhibitors are irreversible?
They bind very tightly, generally using a form of covalent bond, blocking function completely
What happens when enzyme inhibitors are irreversible?
They bind non-covalently and can free dissociate
What are the two types of enzyme inhibition?
NAME?
Where do competitive inhibitors bind?
Active site
What must be true of a competitive inhibitor?
It must also be complementary to the active site
What does a competitive inhibitor reduce?
The proportion of enzyme molecules bound to substrate
Does a competitive inhibitor affect Km or Vmax?
Km
Why does a competitive inhibitor not affect Vmax?
Adding enough substrate will always overcome the effect of the inhibitor
Why does a competitive inhibitor increase Km?
Because the inhibitor competes with the substrate for active sites
Where does a non-competive inhibitor bind?
Another site of the enzyme
Does a non-competitive inhibitor affect Km or Vmax?
Vmax
What does a non-competitive inhibitor decrease?
The turnover number of an enzyme
How does a non-competive inhibitor exert its effect?
It changes the overall shape of the molecule
What other affect can compounds binding outside the active site have?
They can activate the enzyme, rather than inhibit
