Lab 5 Flashcards
The rate of an enzymatic reaction is dependent both on ___ and ____
concentration of the reactants and concentration of the number of enzyme molecules available
The enzyme is a ____. It is not _____
catalyst. It is not destroyed.
When is the rate of the enzyme first measured?
as soon as the substrate and enzyme are mixed for a specified time
The kinetic properties of an enzyme are studied by determining the ______
initial velocity
At the initial velocity, what can you say about:
1. The concentration of the product
2. The backward reaction
- The concentration of the product is very small
- The back reaction
es<—-p
can be ignored
What are enzyme inhibitors?
molecules that inhibit enzyme activity
How do enzyme inhibitors work?
Bind to the enzyme and diminish the velocity of the enzymatic reaction
How are enzyme inhibitors classified?
the type of inhibition
What are irreversible inhibitors?
Irreversibly bind to and inhibit the enzyme. COVALENTLY bound and do not dissociate
What are reversible inhibitors?
Inactivates an enzyme through non covalent, more easily reversed, reactions.
What is Ki?
reversible inhibitor binding constant
How are reversible inhibitors classified?
By their enzyme kinetics
What are the 3 types of reversible inhibitors?
- Competitive Inhibitor
- Noncompetitive inhibitor
- Uncompetitive inhibitor
What is a competitive inhibitor?
Binds to the ACTIVE SITE to which the substrate binds and prevents substrate binding
How is a competitive inhibitor reversed?
By increasing the substrate
What is a noncompetitive inhibitor?
Binds to free enzyme or enzyme complex and prevents enzyme activity
What can you say about reversing a noncompetitive inhibitor?
independent of substrate concentration
What is an uncompetitive inhibitor?
Binds only to ES complex. Prevents release of S and production of P
When does enzyme inactivation occur?
When pH and temperature denature enzymes, and enzyme structure is changed
The order of an enzymatic reaction is related to….
- The number of molecular species
- The rate of the reaction
For a first order enzymatic reaction….
The enzyme concentration is maintained constant. Thus, the enzyme activity is solely dependent on the substrate concentration
When the substrate is much LESS than the Km, the reaction is….
first order with respect to substrate, as the rate is proportional to a single reactant (substrate)
When the substrate is much GREATER than the Km, the reaction is….
zero order with respect to substrate. Velocity is independent of substrate concentration
Initial velocity (V0) is measured when…
the substrate is much greater than the concentration of the enzyme
Initial velocity is measured using….
a constant concentration of enzyme
At a relatively low concentration of substrate, the initial velocity is….
highly dependent on substrate concentration
As the substrate concentration increases, what happens to initial velocity?
increases linearly and rapidly (more enzyme binds to substrate)
What happens when enzymes are increasingly saturated with substrate?
initial velocity changes more slowly
At Vmax, what happens when more substrate is added?
NOTHING - all enzymes are already saturated with substrate. Initial velocity remains constant
When a reaction continues at the maximum initial velocity, what is reached?
a STEADY STATE
An enzyme-catalyzed reaction can roughly be divided into 3 stages:
- enzyme-substrate binding
- Catalysis
- Product release
Michaelis-Menten plot is __vs___
initial velocity vs substrate concentration
The michaelis-Menten plot says that the velocity of an enzymatic reaction is greatest when…
all enzyme’s catalytic sites are filled with substrate
Michaelis-Menten sonstant (km) describes the affinity of….
the substrate to an enzyme
What is km?
the concentration of the substrate that will produce half the vmax
Standard curve shows the relationship between…
absorbance and concentration
The “trend line” of a standard curve is used to….
calculate unknown concentrations
What enzyme is used to catalyze the hydrolysis of p-Nitrophenyl phosphate to p-nitrophenol?
acid phosphatase
In terms of structure, what does acid phosphatase do to p-Nitrophenyl phosphate?
removes the phosphate group and replaces with OH
p-nitrophenol absorbs maximally at…
405nm
Highkm = ___ affinity
low
