Enzyme Kinetics Flashcards
Equation for reaction rate and units of k for 1 and second order reaction
V=K[reactants]
K= 1/s
K=1/MS
What does a pseudo first order reaction mean
One of the reactants is much lower concentration than than the other so the reaction rate doesn’t depend on the lower one
A»»B
A is only included in calculation
What is a zero order reaction
The reaction doesn’t depend on the concentration of reactants
What is initial velocity
How does it get simplified
The moles of product formed per second
The early measurement when time is around 0 and product is around 0
Because of this we can assume [products] is negligible and take it out of the equation to simplify
What happens to initial velocity when conenctrstion of substrate increses
The intially velocity is higher
What is the value of Km
K-1 + k2 / k1
Or the x axis value at Vmax/2
What does the michealis mean ten plot assume
What does this mean
Assume steady state kinetics
The initial velocity is measured while the [ES] (enzyme substrate complex) is relatively stable (constant, meaning the rate of the ES formation = rate of ES breakdown)
What is maximum velocity
Measured When all the enzyme is bound to the substrate so that [ES] = [E] total
It’s directly dependent on enzyme concentration
Vmax = K2 • [E]t
On the part of the graph where [S] is much less than Km what happens to initial velocity
V0= Vmax([S]/Km)
The initial velocity becomes directly proportional to the [S]
On the part of the graph where [S] is much greater than Km what happens
V0= Vmax
The rate is zero order and independent of [S]
On the part of the graph where [S] is equal to Km what happens
V0 = Vmax / 2
km is equal to [S] when the reaction rate is at 1/2vmax
What is Km and what does it depend on
The concentration of substrate where half the enzymes molecules are bound to the substrate
Depends on the type of substrate, ph, temp, ionic strength, type of enzyme
The cellular concentration of a substrate is usually close to what
How does this help
The Km value
This is the point where we have a significant velocity and any small changes up and down in substrate concentration will change the velocity significantly
Sensitive area
What are the two ways to determine km and Vmax
Nonlinear regression analysis
Lineweaver burk plot
What is a non linear regression analysis
You do an experiment an make the michealis menten plot, then put is into a computer system with the equation that the plot gave you
The computer tests values of km and v max that will make the calculated data best fit the experimental data
Fastest and accurate way to find km and Vmax
What is a lineweaver burk plot
Take the inverse of the menten equation (v0= Vmax (s/s+km))
This gives equation in the form of
y=mx + b
The x intercept is -1/km
Y int is 1/vmax
Can find the Vmax and km from the
X and y intercepts
Less reliable because taking reciprocal of larger values
What is the turnover number
Kcat
The number of substrate molecules that the enzyme can convert to product per unit time ONLY when the enzyme is fully saturated with substrate
For ex. If enzyme has kcat value of 500 per second that means the enzyme can take 500 molecules of the substrate per second and turn them each into product
When under these saturated condition we are operating under Vmax
The k2 in the Vmax equation becomes kcat
Kcat=vmax/[E]t
How do you find the amount of time a reaction takes place for kcat
1/kcat give amount of time for a single reaction
Catalytic efficiency : If we want to include kcat in our v0 equation what do we do
Substitute Vmax in the v=0 equation for kcat • [E]t
Catalytic efficiency : When [S] «< Km what happens
V0= (kcat/km) [S] • [E]t
The concentration of E is about equal to the total e concentration
What is kcat/km
The rate constant for ES formation
Is called the specificity constant and measures the catalytic efficiency because it takes into account the rate of catalysis (kcat) and the nature of the enzyme/substrate interaction (Km)
Shows enzymes preference for substrate, if higher more prefer
What is chemotrypsin
A type of enzyme protease that cleaves next to bulky hydrophobic (nonpolar) side chains
Has a higher kcat/km value for phenylalanine meaning the enzyme is good a turning substrate to product
Kcat/km is always Less than
K1
What happens if the rate of product being formed (kcat) is much greater than the dissociation of ES (k-1)
The kcat/km is close to k1
What is k1
The rate constant for the formation of the enzyme substrate complex
The rate limiting step for how efficient the enzyme is going to be
What is the diffusion controlled rate of encounter
The thing that limits the catalytic efficiency, how fast the enzyme can collide with/encounter the substrate in solution
10^8-10^9 is best
If the kcat/km is close to the rate of encounter the enzyme is kinetically perfect
How do temp affect enzyme activity
The activity increases with higher temps but eventuallyy get denatures and stop working
What is tyrosinase
Example of how temp affect activity
The the temp on a cat is lower the colour of fur is darker since enzyme is working better
If temp is higher the enzyme is denature and white fur happens
What is the optimal ph for pepsin
Chemotrypsin
1.5
8
How does Ph affect enzyme activity
Depending on the ph of where the enzyme is, the side chain amino acid residue in the enzyme with be deperotonated or protonated
The optimal activity would be at the peak of the curve
