Lecture 9 - Measuring and Comparing the Activities of Enzymes Flashcards
What does a progress curve do?
Measures the appearance of product or disappearance of substrate with time
When is initial reaction velocity measured?
At/near time zero
What happens if there is sufficient excess of substrate?
As the amount of enzyme increases, the rate of reaction increases
With fixed amount of enzyme, what occurs when excess substrate is added
Curve increases linearly
What happens to the curve when all active sites become occupied?
Rate of reaction stops increasing, previously linear curve starts to slope.
Zero order kinetics:
Rate does not depend on substrate concentration
First order kinetics:
Rate depends on substrate concentration
What does Vmax equal to?
Maximum velocity possible, when there is infinite substrate.
What does KM equal to?
Substrate concentration at which Velocity = Half Vmax
The V vs [S] curve is described by?
The Michaelis-Menten equation
What is the Michaelis-Menten equation?
What are the assumptions for the Michaelis-Menten equation?
- Product is not converted back to substrate.
- Haldane’s stead state assumption (rate of enzyme substrate complex formation = rate of breakdown) [ES] / Time = 0
- Measuring initial rates of reaction means that the substrate concentration does not significantly change over time.
What is the name of the straight line plot?
Lineweaver-Burk plot.
Plotting ___ and ___ gives the Lineweaver-Burk plot
1/V and 1/[S]
What is on the X-axis on the Lineweaver-Burk plot?
1/[S]
What is on the Y-axis on the Lineweaver-Burk plot?
1/V
Y-intersection = ?
1/Vmax
X-intersection = ?
-1/KM
What is the slope equation?
KM / Vmax
Low KM =
high affinity for that particular substrate.
High KM =
low affinity for that particular substrate.
KM = KD =
dissociation constant for that substrate pair.
How Is binding and catalyses described?
An enzyme, E, converts a single substrate, S, to a single product P that is instantly released.
What do the relative speeds of K1 and K-1 do?
Define how tightly substrate binds
What is the rate of catalysis?
K2
What does K2 relate to?
Energy of activation for the transition state
KM = ? (equation)
k-1 / k1
Is KM the same for each substrate?
No
What does the turnover number kcat show?
Number of substrate molecules converted to product, per enzyme, per unit of time when E is saturated with [S]. Helps to define the activity of one enzyme molecule
The ‘peak’ enzyme should have?
A high kcat and a low KM
Kcat / KM is an overall measure of what?
Enzyme efficiency
